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THE DETECTION OF 
POISONS AND POWERFUL DRUGS 



AUTENRIETH— WARREN 



•I, 



LABORATORY MANUAL 

FOR 

The Detection of Poisons 



AND 



Powerful Drugs 



BY 

DR. WILHELM AUTENRIETH 

PROFESSOR IN THE UNIVERSITY OF FREIBURG i. B. 



AUTHORIZED TRANSLATION 

OF THE 

COMPLETELY REVISED FOURTH GERMAN EDITION 



BY 

WILLIAM H. WARREN, Ph.D. 

PROFESSOR OF CHEMISTRY IN WHBATON COLLEGE 



WITH 25 ILLUSTRATIONS . 



> . 






PHILADELPHIA 

P. BLAKISTON'S SON & CO. 

1012 WALNUT STREET 



THE NEW YORK 

JTILDSN FOUNDATIONS 
^ 1920 L 



Copyright, 1915, by P. Blakiston's Son & Co. 



• » • 






THHtMArLMirRHHIiTOilKtFA 



AUTHOR'S PREFACE 



Additional matter in '^ Detection of Poisons" has made the 
fourth edition considerably larger than the third. The seven 
chapters now comprised in the book have been entirely revised, 
but the first three chapters remain unchanged in arrangement. 
Chapter I treats of poisons volatile with steam. Organic 
poisons, especially the alkaloids, form the subject of Chapter II. 
Hydrastine and veronal, introduced into this chapter for the 
first time, have been incorporated into the Stas-Otto process. 
Chapter III deals with metallic poisons. 

The toxic substances included in Chapter IV find no place in 
the three groups just mentioned. As they seldom appear in 
toxicological examinations, they are of theoretical rather than 
of practical significance. The following members of this group 
have been introduced for the first time, namely, cantharidin, 
cytisine, ergot, papaverine, pilocarpine, saponin substances, 
solanine, thebaine, and the toxalbumins, ricin, abrin and crotin. 

Chapter V has to do with special qualitative and quantitative 
methods such as the quantitative estimation of phosphorus in 
phosphorated oils; the electrolytic detection and estimation of 
arsenic; the biological test for arsenic; the destruction of organic 
matter and detection of arsenic by A. Gautier and G. Locke- 
mann; Karl Th. Momer's estimation of minute quantities of 
arsenic; methods of estimating alkaloids by H. Matthes, H. 
Thoms and A. H. Gordin. This chapter also includes A. J. J. 
Vandevelde's estimation of the toxic action of organic com- 
pounds by means of blood haemolysis. 

Chapter VI takes up the estimation of alkaloids and other 
active principles in raw materials (drugs) and in their prepara- 
tions. Pharmacopoeial as well as other estimations such as 
that of nicotine in tobacco, caffeine in tea, coffee, kola prepara- 
tions, etc., pilocarpine in jaborandum leaves, piperine in pepper. 



vi author's preface 

solanine in potatoes, and theobromine in cacao and its prepara- 
tions have been included. The author has endeavored to treat 
these subjects as thoroughly as possible. 

Chapter Vn describes the methods employed in detecting 
carbon monoxide in blood, in recognizing blood itself in stains 
and in differentiating himian from animal blood. 

The new edition, though more comprehensive than the last 
in its scope, has lost nothing in clearness because of the rear- 
rangement of subject matter. Beginners will probably confine 
their attention to the first three chapters. Students of phar- 
macy will undoubtedly add Chapter VI which deals with drug 
assasdng. The other chapters are designed more especially for 
those who wish to become better acquainted with toxicological 
procedures. 

Descriptions of syntheses of organic drugs such as acetanilide, 
antipyrine, phenacetine, pyramidone, salicylic acid, sulphonal 
and veronal allow the student to review the methods employed 
in connection with laboratory work. Structural formulae of 
alkaloids and their cleavage products have been given only when 
they have been definitely determined or shown to be highly 
probable. By introducing this specific information the author 
hopes to stimulate the student's interest in alkaloidal chemistry 
which has become so important within recent years. 

More advanced students will find in fine print brief state- 
ments about the poisonous action of the better known physiologi- 
cally active substances as well as their distribution in and elimi- 
nation from the human organism. Repeated references to 
larger treatises upon toxicology, especially to R. Robert's 
" Intoxikationen", have been made. Numerous citations of 
literature enable the student to consult original articles for fuller 
information. 

The translation of the third and fourth editions into English 
and Spanish and the proposed translation of the fourth edition 
into Italian indicate that colleagues in other countries have 
favorably received this work. 

WiLHELM AUTENRIETH. 
FiiEiBUiiG IN Baden. 



TRANSLATOR'S PREFACE 



The introduction of new matter and certain rearrangements of 
the text make the fourth edition of "Detection of Poisons" 
quite different from the last. Without exception these changes 
have added to the value of the book not only as a laboratory 
manual for students but as a guide for those wishing to make 
practical use of the procedures described. The translation fol- 
lows the German as closely as is consistent with clearness. As 
in the translation of the third edition, the methods from the 
German Pharmacopoeia remain unchanged. 

Aside from the introduction of a few substances that do not 
appear in the earlier editions and the addition of new methods, 
the general plan of the first three chapters is the same as that 
of the last edition. Believing that the subject of so-called 
normal arsenic in Chapter III is not presented in the German 
edition at sufficient length to do full justice to both sides of the 
question, the translator upK)n his own responsibility has imder- 
taken to give a complete statement of the case with citations of 
the principal authorities. 

Most that is new in the book appears in Chapter V which 
treats of special methods of analysis. In addition to the meth- 
ods given in the German edition, the translator has thought it 
worth while to introduce the quantitative estimation of arsenic 
and antimony by the Gutzeit method as worked out by the late 
Professor Sanger. The procedure is so simple that it may appeal 
to some chemists as a desirable substitute for the more com- 
monly used Marsh-Berzelius test. Otherwise the translation 
has not departed from the German text in any essential way. 

William H. Warren. 

Norton, Massachusetts. 



vu 



CONTENTS 

Pa«b 

Author's Preface iii 

Translator's Preface v 

Introduction i 

CHAPTER I 

Tests for Phosphorus and Other Poisons Volatile with Steam from 

Acid Solution 

Phosphorus 5 

Scherer's test; Mitscherlich's test; Blondlot and Dusart'stest; 
(a) in the Presenius-Neubauer apparatus, (h) in the Hilger- 
Nattermann apparatus; Detection of phosphorous acid; Phos- 
phorus in phosphorated oils; Detection and quantitative 
estimation by the Mitscherlich-Scherer method; Metabolism 
in .phosphorus poisoning. 

Further Examination of the Distillate 

Hydrocyanic Acid 19 

Physiological action; Preliminary test; Detection; Quantitative 
estimation; Detection in presence of potassium ferrocyanide; 
Mercuric cyanide; Mercuric cyanide in presence of potassium 
ferrocyanide. 

Carbolic Acid 26 

Action and fate in the organism; Detection; Quantitative esti- 
mations; I. Gravimetrically; 2. Volimietrically (Beckurts- 
Koppeschaar) ; 3. Volumetrically (J. Messinger-G. Vortmann) ; 
Estimation in urine; Carbolic acid in presence of aniline. 

Chloroform 35 

Behavior in the human organism; Distribution in the cadaver; 
Detection; Quantitative estimation in cadaveric material. 

Chloral Hydrate 38 

Detection; Action and fate in the human organism; Quanti- 
tative estimation in blood and tissues. 

Iodoform 41 

Detection. 

Nitrobenzene 42 

Toxic action; Detection. 

Aniline 44 

Toxic action; Detection. 

Carbon Disulphide 46 

Toxic action; Detection; Quantitative estimation of carbon 
disulphide vapor in air. 

ix 



X CONTENTS 

Pagb 

Ethyl Alcohol 49 

Fate in the human organism; Detection. 
Acetone 51 

Occurrence in urine; Detection; Acetone in presence of ethyl 

alcohol; Detection in urine. 

Bitter Almond Water AND Bbnsaldehyde 53 

Synopsis of Group I (Chapter I) 55 

CHAPTER II 

Detection of those Organic Substances which are not Volatile with 

Steam from Acid Solution 

Stas-Otto process. 59 

A. Examination of Ether Extract of the Aqueous Tartaric 

Acid Solution 59 

PiCROTOXIN 61 

Detection in beer. 

colchicin 64 

Picric Acid 65 

Action and Elimination; Detection. 

ACETANILIDE 68 

Action; Detection; Examination of acetamlide urine. 
Phenacetine 70 

Preparation; Detection. 
Salicylic Acid 72 

Detection; Quantitative estimation; Detection in urine. 
Veronal 75 

Preparation; Physiological action; Detection; In urine. 
Antipyrine 78 

Preparation; Detection; In urine. 
Caffeine 79 

Fate in human metabolism; Detection. 

B. Examination of Ether Extract of the Aqueous Alkaline 

Solution 81 

CONIINE 85 

Nicotine 86 

Physiological action; Reactions, 

Aniline 89 

Veratrine 89 

Preparation of crystalline and water soluble veratriBe; Con- 
stitution; Reactions. 

Strychnine 92 

Physiological action; Dotcction; Detection of strychnine in 
presence of brucine. 

Brucine 96 

Atropine 9^ 

Constitution; Roaotiona. 



CONTENTS XI 

Pagb 

HOMATROPINE lOI 

Cocaine loi 

Constitution; Behavior in the animal organism; Detection. 

Physostigmine 105 

Codeine 106 

Narcotine • 108 

Constitution; Detection. 
Hydrastine 112 

Preparation; Constitution; Reactions. 
Quinine 114 

Constitution; Detection. 

Caffeine 118 

Antipyrine 118 

Detection in urine. 
Pyramidone 119 

Preparation; Behavior in the organism; Detection. 

C. Examination of Ether Extract and of Chloroform Extract 

OF THE Solution Alkaline with Ammonia 

a. Ether Extract 122 

Apomorphine 122 

/3. Chloroform Extract 124 

Preliminary test for morphine; Purification of crude morphine. 

Morphine 126 

Constitution; Detection; Behavior in the animal organism. 

Narceine 131 

Constitution; Reactions. 

Synopsis of Group II tCn after II) 134 

CHAPTER III 

Examination for Metallic Poisons 

Fresenius-v.Babo Method of destroying organic matter .... 141 

Destruction of organic matter with free chloric add 144 

C. Mai's Method of destroying organic matter 145 

Precipitation with hydrogen sulphide 145 

Metallic Poisons I: Examination of that portion of the hydrogen 
sulphide precipitate soluble in ammonia-anmionium stilphide. 

Arsenic 149 

Marsh-Berzelius method; Fresenius-v. Babo method; Betten- 
dorfif's arsenic test; Gutzeit's arsenic test. 

Antimony, Tin, Copper 156 

Metallic Poisons II: Examination of that portion of the hydrogen 

sulphide precipitate insoluble in ammonium sulphide .... 158 

Mercury, Lead, Copper, Bismuth, Cadmium 158 

Metallic Poisons III: Examination for Chromium and Zinc . . . 161 

Zinc 161 



• • 



Xll CONTENTS 

Pagb 

Chromium 162 

Metallic Poisons IV: Examination for Barium, Lead and Silver of 
the insoluble residue left on treatment with potassium chlorate 

and hydxx>chloric acid 163 

Synopsis of Group III (Chapter III) 164 

The Action of Heavy Metals 165 

Pate, Distribution and Elimination of Metals in the body . 166 

CHAPTER IV 

1. Examination for those Poisons which do not belong to the Three 

Main Groups of Poisons 

Mineral Acids 

Hydrochloric Acid . 176 

Nitric Acid 177 

Sulphuric Acid 179 

Sulphurous Acid 181 

Oxalic Acid 182 

Toxic action; Distribution in the organism; Detection. 

Detection of Free Alkalies 

Potassium Hydroxide, Sodium Hydroxide, Ammonia 185 

Potassium Chlorate 187 

Toxic action; Detection; Quantitative estimation; Behavior 

during putrefaction; Detection in meat. 

Examination for Santonin, Sulphonal and Trional 191 

Santonin 191 

Constitution; Behavior in the organism; Detection 
Sulphonal 193 

Preparation; Detection; In urine; Detection of haematopor- 

phyrin in urine. 
Trional 196 

2. Powerful Organic Substances of Rare Occurrence in Toxi- 

COLOGICAL Examinations 

Cantharidin 196 

Constitution; Detection. 
Cytisine 198 

Preparation; Toxic action; Detection. 
Digitalis Bodies 200 

Digitonin, Digitoxin, Digitalinum verum. 
Ergot 202 

Alkaloids; Sclererythrin; Detection of ergot in flour; Detec- 
tion and estimation of the alkaloids. 
Opium 205 

Meconicacid; Meconin; Selenious-Sulphuric acid, a reagent for 

opium alkaloids. 



CONTENTS Xiii 

PilGB 

Papaverine ao8 

Constitution; Detection. 

Pilocarpine 210 

Ptomaines 212 

Saponins 213 

Physiological action; Detection in foaming beverages, such as 

beer, etc.; Detection of githagin in flour. 

Haemolysis and Physiological Salt Solution 216 

SoLANiNE 217 

Toxic action; Detection. 
Tbebaine 220 

Constitution; Detection. 
Toxalbumins 221 

Abrin, Ridn; Crotin; Coagtilation of blood and defibrinated 

blood. 

CHAPTER V 
Special Methods 

Quantitative Estimation of Phosphorus in Phosphorated Oils 224 
I. W. Straub's method; 2. A. Fr&nkel's and C. Stich's method. 

Special Methods for Detecting Arsenic 226 

Separation of arsenic as arsenic trichloride 226 

Electrolytic detection of arsenic 226 

Destruction of organic matter and detection of arsenic by A. 

Gautier and G. Lockemann 227 

Electrolytic estimation of minute quantities of arsenic by C. 

Mai and H. Hurt 230 

Quantitative estimation of arsenic and antimony by the Gutzeit 

method 233 

Biological detection of arsenic by means of penicillium brevi- 

caule 235 

Detection of arsenic in organic arsenic compounds 238 

Cacodylic acid; Arrhenal; Atoxyl; In urine; 

Quantitative estimation of minute quantities of arsenic by Karl 

Th. M6mer 240 

Detection of Salicylic Acid in Foods and Beverages; In Wine, 

Meat Products, Milk 243 

Maltol 244 

Use of Chloral Hydrate in Toxicologic al Analysis by R. Mauch. 

Alkaloidal Estimations 244 

1. By the picrolonate method of H. Matthes 246 

2. By precipitation with potassium bismuthous iodide and de- 
composition of the precipitate with alkali hydroxide-carbonate 

by H. Thoms 248 

3. By the method of H. M. Gordin 250 

Quantitative estimation of strychnine and quinine in presence 

of each other 251 



XIV CONTENTS 

* 

Pagb 
Estimation of the toxicity of chemical compounds by blood 

haemolysis by A. J. J. Vandevelde 251 

CHAPTER VI 

QUANTITATIVB ESTIMATION OF ALKALOIDS AND OTHER POWERFUL SUB- 
STANCES IN Raw Materials and in their Preparations 

Alkaloidal Estimations op Drugs and Their Pharmaceutical Prepa- 
rations According to the German Pharmacopoeia. . . 253 

Estimation of alkaloid in aconite root 254 

Estimation of cantharidin in Spanish fly 256 

Estimation of cinchona alkaloids 257 

I. In cinchona bark; 2. In aqueous extract of cinchona and 
in alcoholic extract of cinchona. 
Estimation of quinine in mixtures of cinchona alkaloids by the 
sulphate method 261 

I. Cinchona bark; 2. Cinchona extract. 
Estimation of colchicin in colchicum seeds and in colchictmi 

corms 262 

Estimation of alkaloid in pomegranate bark 264 

Estimation of caffeine in coffee, tea, kola nuts and Guarana 
paste 264 

I. C. C. Keller's method; 2. A. Hilger-A. Juckenack's 
method. 3. A. Hilger-H. GOckel's method; 4. Socolof- 
Trillich-G6ckel-method; 5. E. Katz's method; 6. K. 
Dieterich's method. 

Estimation of alkaloid in ipecacuanha root 270 

Estimation of nicotine in tobacco 272 

I. R. Kissling's method; 2. C. C. Keller's method; 3. J. 
Toth's method. 

Estimation of hydrastine in hydrastis extract 274 

Estimation of bcrberine 27$ 

Estimation of hydrastine by the picrolonate method of H. 
Matthes and O. Rammstedt 275 

I. In fluid extract of hydrastis; 2. In hydrastis root. 
Estimation of morphine in opitun and in its pharmaceutical 
preparations 276 

I. In opixim; 2. In extract of opium; 3. In wine of opium 
and in tincture of opium. 
Estimation of pilocarpine in jaborand tun leaves 279 

I. G. Fromme's method; 2. H. Matthes and O. Ramm- 
stedt's method. 
Piperine and its estimation in pepper 281 

I. J. Kdnig's method; 2. Cazeneuve and Caillot's method. 
Estimation of santonin in wormseed 2g2 

I. K. Thater's method; 2. J. Katz's method. 
Estimation of solanine in potatoes 23^ 



CONTENTS XV 

Page 
I. O. Schmiedeberg and G. Meyer's method; i. F. v. Mor- 
genstem's method. 
Estimation of alkaloid in nux vomica and its preparations . . 286 

C. C. Keller's method 286 

Method of the German Pharmacopoeia 287 

I. In nux vomica; 2. In extract of nux vomica; 3. In 
tincture of nux vomica. 

H. Matthes and O. Rammstedt's method 289 

I. In extract of nux vomica; 2. In tincture of nux vomica; 
3. In nux vomica. 
Estimation of strychnine in mixtures of strychnine and brucine 

by C. C. Kellei--H. M. Gordin 291 

Estimation of theobromine and caffeine in cacao and in choco- 
late ^ 291 

Estimation of alkaloid in the leaves of atropa belladonna, hyo- 

scyamus niger and datura strammonitmi 293 

Estimation of alkaloid in extract of belladonna, according to 

the German Pharmacopoeia, in extract of hyoscyamus . . . 294 

Assay of officinal extracts by E. Merck 295 

Extract of belladonna; Extract of cinchona; Extract of 
strychnine. 

CHAPTER VII 

Detection of Carbon Monoxide Blood, Blood Stains and Human 

Blood 

1. Recognition of carbon monoxide blood 297 

2. Detection of blood stains 300 

Hsematin 301 

Spectroscopic detection of blood 303 

Other tests for blood 305 

Schdnbein-van Deen's test; Vitali's procedure in this 
test; Schaer's procedure; Aloin test. 

3. Biological detection of human blood 307 

APPENDIX 
Preparation of Reagents 

A. General alkaloidal reagents 310 

B. Special reagents and solutions 313 

C. The indicator iodeosine 315 

Index 317 



INTRODUCTION 



Nearly all the common poisons and drugs may be placed in 
one of three groups. This classification, based upon the 
chemical behavior of these substances during isolation from mix- 
tures is as follows: 

Group I. — The members of this group volatilize without 
decomposition when heated and distil from an acid solution with 
steam. Yellow phosphorus, hydrocyanic add, carbolic add, 
chloroform, chloral hydrate, iodoform, aniline, nitrobenzene, 
carbon disulphide and alcohol are the prindpal substances of 
this dass. 

Group II. — The members of this group are non-volatile, 
organic substances which do not distil from an add solution 
with steam. But hot alcohol containing tartaric acid will 
extract them from extraneous matter. Alkaloids, many glu- 
cosides and bitter prindples, as well as certain synthetic organic 
drugs like acetanilide, phenacetine, antipyrine, pyramidone, 
sulphonal and veronal comprise this group. 

Group III. — This group includes all poisonous metals. 

In toxicological analysis, therefore, poisons are divided into 
three groups, each of which has its own spedal methods of pro- 
cedure. A few poisons like mineral acids, caustic alkalies, 
oxalic add and potassium chlorate cannot be conveniently 
placed in these three groups owing to differences in solubility 
and other peculiarities. Spedal tests for such substances 
must be made with a separate portion of material. Chapter IV 
contains a description of the methods used in identifying these 
substances. 

The material must be thoroughly mixed and divided into 

three or four approximately equal portions, unless the analysis 

is to be limited to the detection of a single well-defined substance. 

One portion is tested for non- volatile, organic substances (Chap- 

1 



2 INTRODUCTION 

ter II). The second portion is examined for volatile poisons 
(Chapter I) and the residue from this portion is used in testing 
for poisonous metals (Chapter III). The third portion is 
tested for substances considered in Chapter IV. The fourth 
portion is held in reserve in case additional material is needed 
to verify a doubtful result, or to replace a portion accidentally 
lost during analysis. 

Occasionally it is advisable to depart from the general pro- 
cedure and follow a special method, especially in detecting a 
single poison, or in estimating it quantitatively. For instance, 
pure ether would not be the best solvent to use in extracting 
strychnine quantitatively from an alkaline solution. A mix- 
ture of ether and chloroform, or better pure chloroform would 
be preferable, since strychnine is more soluble in the latter 
solvent than in pure ether. For the same reason chloroform 
should be used in the quantitative extraction of caffeine or 
antipyrine. When only a small quantity of material is available 
for analysis, tests for all three groups of poisons may be made 
with the same portion. In this case after removal of volatile 
poisons (Chapter I) the residue should be divided into two un- 
equal portions. The larger portion should be tested for non- 
volatile, organic poisons (Chapter II). The smaller portion 
together with the residue left after extracting non-volatile, or- 
ganic poisons should be tested for poisonous metals (Chapter 
III) . It is advisable, however, even in such a case to reserve a 
portion of material for any contingencies. 

Organs of the human body like liver, kidneys, spleen, heart, 
brain, stomach or intestines with contents should be cut into 
small pieces and then finely chopped before being examined 
chemically. An organ should first be cut into small pieces with 
sharp, clean scissors and then minced with a clean chopping 
knife in a new wooden bowl, or a small meat machiney which 
has been carefully cleaned, may be used. Material may be 
held with nickel plated tongs while being cut with scissors. 



DETECTION OF POISONS 



CHAPTER I 



VOLATILE POISONS 



Yellow Phosphorus and Other Poisons Volatile from Acid Solution 

with Steam 

Scherer's Test. — This test should precede the distillation 
described on page i8. The principle of the test is that moist 
phosphorus vapor and silver nitrate form black silver phosphide 
(AgsP), metallic silver, phosphoric and sometimes phosphorous 
add. Place the finely divided material in a small flask and 
cover with water if a sufiicient quantity is 
not present. Cut a V-shaped slit in the 
cork and place the latter loosely in the 
mouth of the flask so that the two strips of 
filter paper are freely suspended (Fig. i). 
Moisten one strip with silver nitrate and 
the other with lead acetate solution.^ 
Warm gently upon the water-bath (40 to 
50°).^ If the silver paper is darkened but 
not the lead paper, yellow phosphorus may 
be present. If both papers are darkened, 
hydrogen sulphide also is present. In the 
latter case yellow phosphorus may be pres- 
ent with hydrogen sulphide. In absence of 
hydrogen sulphide, darkening of the silver 
paper is not final proof of yellow phosphorus, for any volatile 
organic substance having reducing properties, as formalde- 

^ A more sensitive "lead paper" may be obtained by using alkaline lead solu- 
tion prepared by adding excess of sodium hydroxide to the solution of a lead 
salt whereby Pb(OH)(ONa) and Pb(ONa)i are formed. 

' Temperatures in this book are expressed in Centigrade degrees. Tr. 

3 




Fig. I. 



*. 



- ». 



4 DETECTION OP POISONS 

hyde (H.CHO), or formic add (H.COOH), may give the 
same result. 

Scherer's test is of value in proving the absence rather than the presence of 
yellow phosphorus. It is a good prdiminary test, as it excludes phosphorus if 
the silver paper is unchanged. 

Distillation. — Place a portion of finely divided and thoroughly 
mixed material in a large round-bottom flask and add enough 
distilled water for free distillation. Then add tartaric add 
solution drop by drop until the mixture is add after thorough 
shaking. Practice analyses^ usually require 20 to 30 drc^ of 
10 per cent, tartaric add solution. 

In examining animal material, as the stomach or intestines 
and contents, or organs, like liver, spleen and kidneys, it is often 
unnecessary to add much water because enough is usually pres- 
ent. First chop the material in a wooden tray with a sted knife. 
In a medico-legal analysis the tray should be new. A meat 
machine which has been carefully deaned may be used. Thin 
the material with a littie distilled water, addify with dilute 
tartaric or sulphuric add and finally distil. 

If Scherer's test is positive, begin distilling with the Mit- 
achcrlich apparatus (Fig. 2) ; but if negative, distil in the usual 
way with the Liebig condenser (see page 18). The distillate 
may contain: 

Yellow phosphorus mtrobenzene 

Hydrocyanic add Aniline 

Carbolic acid Ethyl alcohol 

Chloroform Acetone 

Chloral hydrate Carbon disulphide 

lodofonn Benzaldehyde 

Bitter almond water 

* Laboratory practice in detecting poisons may be given by mixing small quui- 
titiea (from 0.03 to 0.05 or o.i gram) of a poison witk dry bread or biflcidt 
crumbs, meal or meat. Finely chopped organs (liver, " Iddney, spleen, etc.), 
sausage meat, beer, wine or milk may be used. Drugs Hke morphine, codeine, 
quinine, acetanilide, phenacetine, antipyrine, cafiFein^**<antonin, sulphonal, 
veronal, calomel, tartar emetic, subnitrate of bismuth, etc., may be mixed with 
powdered cane- or milk-sugar. The last tikd of practice is especially suitable 
for students of pharmacy. 






VOLATILE POISONS 



TELLOW PHOSPHORUS 
Mitscherlich Method of Detecting Yellow Phosphorus 

The principle of this method is that yellow phosphorus is 
volatile with steam and becomes luminous in contact with air. 
The phosphorescence is best seen in a dark room. 








Fig. 2. — Mitscherlich Apparatus. 

Procedure. — Arrange the apparatus as in Fig. 2. Support 
the condenser in a vertical position and connect the upper end 
with the flask by a glass tube about 8 mm. internal diameter. 
This tube has two right-angle bends and each end passes through 



6 DETECTION OF POISONS 

a cork. li'dva condenser and tube scrupulously clean to avoid 
interference with the phosphorescence. 

Have the flask at most not more than a third full. This pre- 
caution is necessary because many materials, containing protein 
substances like albumin, albumose, etc., and starchy matter, 
when distilled in aqueous solution, cause more or less foaming 
which is liable to carry over s^jlid matter into the receiver. Use 
as the receiver an Jtrlenmeyer flask containing a little distilled 
water (^i to 5 cc.j into which the end of the condenser dips. 
This precaution prevents loss of easily volatile substances like 
hydrocyanic acid and chloroform. Heat the flask upon a wire 
gauze of fine mesh, asbestos plate or sand bath and bring the 
contents to boiling by raising the temperature gradually. 
There is some danger of burning or carbonizing organic matter 
on the bottom of the flask, if heat is applied too strongly or 
rapidly. When boiling begins, make the room as dark as pos- 
sible and watch for phosphorescence in the tube and condenser. 
It usually appears as a luminous ring or band in the upper part 
of the condenser. When this is distinctly visible, the presence 
of yellow jihosjihorus is established. Phosphorescence during 
distillation with steam is very characteristic of yellow phos- 
phorus and frequently is the only sure and unquestionable test 
for this element. 

Phosphorescence is a process of oxidation by which phos- 
phorus vapor is changed to phosphorous acid. Should it not 
appear immediately, distillation must be continued for some 
time, since certain substances like ethyl alcohol, ether, turpen- 
tine and many other ethereal oils either prevent the phenomenon 
entirely or seriously retard it. Considerable carbolic acid, 
creosote, chloroform, chloral hydrate, as well as hydrogen 
sulphide, may completely prevent phosphorescence. 

K. Polstorfl and J. Mcnsching^ have shown that mercuric 
cl^loridc as well as other mercury compounds may also interfere 
with phosphorescence. Possibly mercuric chloride carried over 
by $team, is reduced to metallic mercury by phosphorus vapor. 
In chat case the metal should appear in the distillate. The 

^Btaichte der Deutschen chcmischen Gcsellschaft 19, 1763 (1886). 



VOLATILE POISONS 7 

fact that both metallic mercury and phosphoric acid can be 
detected in the distillate favors the supposition that action 
takes place between phosphorus vapor and mercuric chloride. 

Phosphorescence, however, often appears when these sub- 
stances have passed over. But even when 'prolonged dis- 
tillation fails to give a positive result, this must not be accepted 
as final proof of the absence of phosphorus until other tests 
have been made. Whatever the result, evaporate a portion 
of the distillate to dryness on the water-bath with excess of 
saturated chlorine water, or with a little fuming nitric acid. 
Phosphorus always imparts a strong odor to the distillate. 
Small drops of phosphorus appear if the quantity is large, and 
the solution contains phosphorous acid. Dissolve the residue 
from evaporation in 2 to 3 cc. of water and test in two sepa- 
rate portions for phosphoric acid. 

1. Ammonium Molybdate Test. — Acidify the solution with 
a few drops of concentrated nitric acid. Add an equal volume 
of ammonium molybdate solution 
and warm to about 40°. Phos- 
phoric acid precipitates yellow 
ammonium phospho-molybdate. 

2. Ammonium Magnesium 
Phosphate Test — Add magnesia 
mixture^ to the second portion. 
Phosphoric add gives a white 
crystalline precipitate of ammo- 
nium magnesium phosphate (H4N) 

ikr Tkr^ ^Tx /-w tr« i i • FiG. $, — Ammonium magiiesium 

MgP04.6H20. Vigorous shakmg phosphate crystals. 

favors precipitation. When only 

traces of phosphoric acid are present, long standing is necessary 
before the precipitate appears. Always examine the precipi- 
tate with the microscope. It should consist of well-formed 

^Magnesia mixture is a clear solution prepared by mixing equal volumes of 
magnesium chloride, ammonium chloride and ammonium hydroxide (about 
10 per cent.) solutions. It contains the readily soluble double chloride of ammo- 
nium and magnesium which is not decomposed by ammonium hydroxide. This 
reagent is prepared as needed and should be perfectly clear and colorless. 




8 DETECTION OF POISONS 

crystals or at least should be crystalline. These crystals are 
transparent, adcular prisms (Fig. 3). 

Notes. — A. Fischer^ states that substances interfering more or less with the 
detection of phosphorus by the Mitscherlich method are usually less troublesome 
if Hilger and Nattermann's procedure is used (see page 15). The essential 
feature of the latter process consists in allowing steam charged with phosphorus 
to pass into the air, or in admitting air into the apparatus. 

Detection of Phosphorus and Phosphorous Add 

(Blondlot*-Dusart«) 

When the Mitscherlich method fails to show phosphorus, it 
is often necessary to test for phosphorous acid. This is the 
first product in the oxidation of phosphorus and is easily formed. 
The Blondlot-Dusart method shows the slightest trace of phos- 
phorous (HsPOs) and hypophosphorous (HsPOj) adds as well 
as yellow phosphorus. The method consists in converting 
yellow phosphorus into phosphine (PHj) by nascent hydrogen. 
The lower oxidation products of phosphorus, namely, hypo- 
phosphorous (HsPOi) and phosphorous (HsPOs) adds,* when 
warmed with zinc and dilute sulphuric add are reduced to 
phosphine by nascent hydrogen: ' 

H,PO, -f 4H - PH, -f 2H,0, 
H.PO. + 6H " PH, + 3H,0. 

Phosphine, or hydrogen charged with phosphorus vapor, 
burns with a characteristic green flame (Dusart's reaction) : 

2PH, + 4O, - P,0, + 3H,0. 

The green flame is easily recognized by depressing a cold 
porcelain dish or plate upon the flame. Detection of phosphorus 
by the BIondlot-Dusart method depends upon these two facts. 

A toxicological analysis usually deals with the detection of 
traces of yellow phosphorus. Hydrogen after acting in the 
nascent state upon the material is not directly examined for 

* Pfluegcr's Archiv 07, 578 (igo^). 

•Journal de Phnrmadc et cir Chimic (,0, 4 \ aS* 
•Comptes rcndus dr PAnidcmir dcs Sciences, 43. 1126, 

* Nascent hydrogen will not reduce ordinary, or ortho-phosphoric add (H»PO«), 
and its derivatives, pyrophosphoric OUP1O7) and meta-phosphoric (HPOj) adds, 
to phosphine. 



VOLATILE POISONS 9 

phosphorus but is first passed into dilute silver nitrate solution. 
Phosphorus and phosphine precipitate black silver phosphide^ 

(Ag,P) : 

PH, + 3AgN0, - Ag,P + 3HNO,. 

Thus traces of yellow phosphorus may be concentrated in the 

silver precipitate from which nascent hydrogen will liberate 

phosphine: 

Ag,P + 3H - PHi + 3Ag. 

If hydrogen produces a black or gray precipitate in the silver 
solution, phosphorus is not necessarily present, as hydrogen 
sulphide, arsine, stibine and reducing organic compounds be- 
have similarly with silver nitrate. A black precipitate there- 
fore should always be examined for phosphorus by the Dusart 
reaction. 

In the detection of yellow phosphorus, the Blondlot-Dusart 
method combines two distinct operations, namely: 

1. Preparation of the silver phosphide precipitate. 

2. Examination of this precipitate in the Dusart apparatus. 
Procedure, i. Preparation of Silver Phosphide. — Thin the 

finely divided material with water in a capacious flask where 
hydrogen is being evolved from phosphorus-free zinc and pure 
dilute sulphuric add (1:5). In testing for phosphorous add 
alone (see page 14) use the filtrate from an aqueous extract 
of the material, or the filtrate from the residue left after the 
Mitscherlich distillation (see page 5). Nascent hydrogen 
should act for 1.5 to 2 hours, or even longer, and pass through 
neutral silver nitrate solution in the receiver at the end of the 
apparatus. If yellow phosphorus is present, the hydrogen will 
contain phosphorus and phosphine and cause a black predpi- 
tate of silver phosphide in the silver nitrate solution. Collect 
the predpitate upon a small ash-free paper, wash with a little 
cold water and examine in the Dusart apparatus as described 
elsewhere. 

^ Phosphorus cannot be determined quantitatively as silver phosphide because 
this compound is partially decomposed by water. Phosphoric and phosphorous 
adds pass into solution: 

(a) 2Ag,P 4- sO + 3H1O = 6Ag -f 2H,P0«, 

(b) 2Ag,P -h 3O -h 3H2O = 6Ag -h 2Ha*0,. 



10 



DETECTION OF POISONS 



If there is silver phosphide in the precipitate, the filtrate will 
contain phosphoric or phosphorous add (see Note, page 9). 
To detect phosphoric acid, first add hydrochloric acid to remove 
excess of silver from this filtrate. Filter through paper pre- 
viously well washed with add and water and completely expel 
hydrochloric add from the filtrate by evaporation upon the 
water bath with concentrated nitric add. Dissolve the resi- 
due in a little warm water and test for phosphoric add with am- 
monium molybdate or magnesia mixture. 

2. Examination of the Silver Precq)itate (AgsP) for Phos- 
phorus. — Two forms of apparatus may be used for this purpose, 
namely: 

(a) Fresenius-Neubauer^ .^yparatus. — Generate hydrogen 
in flask A (Fig. 4) from pure phosphorus-free zinc and dilute 




Kio» 4. Krt^cwlvw-Ncub*ucr .\|>(>armtus. 



sulphuric aciil. Kill U-tubo C" with pieces of pumice stone satu- 
ratiHl with coiumt rated potassium hydroxide solution to ab- 
sorb ai\y hyilrv^gon sulphide, Tse harvi glass for tube D and 

> V\ R. Ftt»scn\U!». Vjunlitrttivc chrmw^ht* Am^l^N-tf^ XVI edilioii. p«ce 521, 



VOLATILE POISONS 1 1 

have the tip F of platinum. The part marked E^ is a glass 
stop-cock or screw-tap. Reservoir B serves to hold liquid from 
A when cock E is closed. A platinum tip is essential, other- 
wise the flame instead of being colorless will always be yellow 
from sodium in the glass. The place where the platinum tip is 
fused into the glass should be cooled by wrapping cotton around 
the glass and keeping it moist. 

Procedure. — Open E and let hydrogen from A pass for some 
time through the apparatus to expel air. Then close E and 
liquid in A will rise into B. Now open E just enough to allow 
hydrogen to bum with a small flame which should be colorless 
in the dark. If there is no trace of green in the inner cone and a 
porcelain dish depressed upon the flame does not show an em- 
erald green coloration, hydrogen is phosphorus-free. It is well 
to repeat this test. To test the silver precipitate for phosphorus, 
wash it with the paper into B with a little water. 

When the entire precipitate is in A, close E until all the liquid 
has risen from A into B. Then open E, light the hydrogen and 
examine the flame in the dark. If the precipitate contains a 
trace of silver phosphide, the inner cone will be green and a 
porcelain dish depressed upon the flame will show an emerald 
green coloration. Have the hydrogen flame small so that its 
color may be observed for some time. 

(6) Hilger-Nattermann^ Apparatus. — Reduction takes place 
in a IOC cc. flask closed by a rubber stopper with three holes, 
two of which are for right- angle tubes just passing through the 
stopper and the third for a thistle tube going to the bottom of 
the flask (Fig. s) . Hydrogen from a Kipp generator enters the 
flask by one tube and leaves by the other. Attach to the latter 
a U-tube filled with pieces of pumice stone saturated with 
concentrated potassium hydroxide solution to absorb hydrogen 
sulphide. Connect the other end of the U-tube with a hard 



^ Fresenius and Neubauer use a screw pinch-cock instead of a gas-cock but by 
means of a short rubber connector they interpose an ordinary cock between the 
gas flask A and the U-tube C. 

'Forschungsbericht fiber Lebensmittel und ihre Beziehungen zur H3rgiene, 
etc., 4, 241-258 (1897). 



12 



DETECTION OF POISONS 



glass tube tipped with platinum.^ Cut the paper containing 
the precipitate into small pieces and place in the flask which 
contains in addition a few pieces of phosphorus-free zinc and 
enough water to seal the thistle tube. Light the hydrogen after 
it has passed through the apparatus for some time and been 
found free from air by the usual test. Seen in the dark the 
flame should be entirely colorless and burn without a green cone 









=^ 



<r=^ 



'^^=^ rr=^ 








Fig. 5. — ^Hilger-Nattennan Apparatus. 

or a greenish glow.^ Hilger and Nattermann advise a spectro- 
scopic examination of the flame to determine the purity of the 
zinc. Pure zinc gives a hydrogen spectrum which shows only 
an orange colored line in place of the yellow sodium line. The 
minutest trace of phosphorus will give three green lines lying 
to the right of the line D. The color of two of these lines is more 
pronounced than that of the third. Having thus tested the 
purity of zinc and sulphuric add, pour a few cc. of dilute sul- 
phuric acid (1:5) through the thistle tube into the flask con- 
taining zinc and the silver precipitate. If the latter contains 
phosphorus, the flame will show, though not always at once, a 
green coloration which should be examined with the spectro- 
scope. 

^ Hilger and Nattermann use a platinum tipped blow-pipe instead of a glass 
tube tipped with the same metal. Cotton, which is kept moist and acts as 
a cooler, is wrapped around the blow-pipe below the tip. 

'Zinc entirely free from phosphorus which will stand this test is difficult 
to obtain. 



VOLATILE POISONS 13 

The Mitscherlich method affords a distillate especially suit- 
able for the Blondlot-Dusart test. If this imparts a green color 
to the hydrogen flame, there can be no question about the pres- 
ence of phosphorus. 

Although the Blondlot-Dusart test is very delicate, many 
chemists refuse to accept it as a substitute for the Mitscherlich 
test. Selmi states that animal material like brain, which con- 
tains organic phosphorus compounds, yields after putrefaction 
a distillate that often gives a black precipitate with silver ni- 
trate solution. This will impart a green tinge to the hydrogen 
flame in the Blondlot-Dusart test. 



Z. HaUsz,^ however, has failed to confirm Selmi's results. He examined two 
kinds of animal material by the Blondlot-Dusart method. First, he tested nor- 
mal brains (man, calf, hog); second, the brain and other organs of rabbits that 
had been given poisonous doses of phosphorus by the mouth and subcutaneously. 
He examined these organs when fresh and also from week to week after more or 
less pronounced putrefaction had set in, but could not detect phosphorus in the 
brain in a single instance. These experiments disprove the earlier idea that 
phosphorus normally present in the brain is so changed during putrefaction that 
it can be detected by the Blondlot-Dusart reaction. He also failed to detect 
phosphorus in the brain of rabbits poisoned by thb element, though he found it in 
other organs, as stomach and intestines, and in those rich in blood, as liver, 
limgs and kidneys. He could always detect small or large quantities of phos- 
phorus in any organ which this element had directly reached, or by which it had 
been indirectly absorbed. If any compound containing phosphorus is really 
formed in the brain during putrefaction, Hal&sz concluded that it is not volatile 
with steam and does not give the Blondlot-Dusart reaction. On the basis of these 
experiments Hal&sz holds that the Blondlot-Dusart method of detecting phos- 
phorus is just as reliable for forensic purposes as that of Mitscherlich. 

Procedure of Halasz in the Blondlot-Dusart Method 

Make a thin mixtiure of very finely divided material and boiled water in a flat- 
bottom flask where hydrogen is being generated from phosphorus-free zinc and 
dilute sulphuric add. Warm upon the water-bath and pass the gas through an 
absorption tube provided with several bulbs and containing neutral silver nitrate 
solution. Concentrated sulphiuic acid and a little platinic chloride may be added 
toward the end to hasten the evolution of gas. Nascent hydrogen thus acts upon 
phosphorus in the animal material for 2-2.5 hours. Finally wash the silver 
precipitate carefully with water and transfer it with the paper to the Blondlot- 
Dusart apparatus. 

^Zeitschrift fttr anorganische Chemie 26, 438 (1901). 



14 DETECTION OF POISONS 

Detection of Phosphoroas Add 

The reduction of phosphorous add to phosphine by zinc and dilute sulphuric add 
takes place very slowly. Hilger and Nattermann state that even a few milli- 
grams require the action of nascent hydrogen for lo to 14 days. Moreover care- 
ful manipulation is necessary because silver phosphide is quite imstable. Water 
decomposes thb substance into metallic silver and phosphorous add and the 
nitric add present oxidizes the latter to phosphoric add. Therefore when spedal 
attention must be given to phosphorous acid, Hilger and Nattermann recommend 
examining the silver predpitate (presumably AgiP) after 2 days, or at most 
3, for phosphorus by the Blondlot-Dusart method and the filtrate for phos- 
phoric acid (see page zo). 

Detection of Phosphorus in Phosphorated Oils 

1. Straub's^ Test — If a phosphorated oil is placed on the 
surface of copper sulphate solution, phosphorus will gradually 
pass from the oil to the aqueous solution and first form black 
copper phosphide. The latter, acting as a carrier of oxygen, 
oxidizes phosphorus still in the oil to phosphoric acid which 
dissolves in the water. 

Shake 10 cc. of phosphorated oil in a test-tube with 5 cc. of i 
per cent, copper sulphate solution. According to the amount of 
phosphorus a black or light brown coloration will appear in the 
emulsion at once, or in a few minutes, and at most after 2 
hours. Phosphoric add in the aqueous solution may be recog- 
nized by ammonium molybdate. At least 0.0025 P^^ cent, of 
phosphorus may be detected in this way. 

A practical, therapeutic application of this reaction may be made in acute phos- 
phorus poisoning. Administration of copper sulphate solution may prevent 
absorption of free phosphorus still in the gastro-intestinal tract. 

2. The Mitscherlich test is also applicable to phosphorated 
oils, even though the oil may contain only 0.0002 gram of 
phosphorus in 100 grams. But phosphorescence will not 
appear unless air is admitted into the tube from time to time. 

Phosphorus in oils cannot be determined quantitatively by 
the distillation method, for not more than 36 to 41 per cent, of 

^W. Straub, Mtinchener medizinische Wochenschrift 50, 1145; Archiv der 
Pharmazie 241, 335 (1903); and Zeitschrift fttr anorganische Chemie 35, 460 

(1903). 



VOLATILE POISONS 15 

phosphorus will distil over. The quantitative method recom- 
mended by Straub (see page 224) may be used in that case. 

Detection and Quontititlve Bstinuition of Phosphorus 
(MitscherUch-Scherer) 

Acidify a weighed portion of material with dilute sulphuric 
add and add a little ferrous sulphate. Distil in a gentie stream 
of carbon dioxide, using a large flask fitted with a two-hole 
stopper. Expel air completely from the apparatus by carbon 
dioxide before heating. Use as receiver a flask fitted with a 




T-NattennauQ ApparatuB (or Detecting and Quantit&tivdy Esti- 
mating Phosphorus. 

two-hole stopper. Pass the end of the condenser through one 
hole and connect the other with a U-tube containing silver 
nitrate solution. Evaporate the distillate upon the water 
bath with strong bromine water, or with concentrated nitric 
add, to oxidize phosphorus or any phosphorous add formed. 
Dissolve the residue in a little water and predpitate phos- 



16 DETECTION OP POISONS 

phoric add with magnesia mixture. Weigh the precipitate as 
magnesium pyrophosphate, MgsPsOr. Heat the contents of 
the U-tube with concentrated nitric add. Predpitate silver 
as silver chloride and filter. Concentrate the filtrate by evapo- 
ration and predpitate phosphoric add with magnesia mixture 
as before. Combine this predpitate with the other. In dis- 
tillation some phosphorus separates as globules in the first 
recdver and any carried over as vapor by carbon dioxide is re- 
tained by silver nitrate solution. As the steam distillation of 
phosphorus is very slow, the process should be carried on for at 
least 3 hours. Hilger and Nattermann recommend the 
apparatus in Fig. 6 not only for detecting phosphorus but for 
estimating it quantitativdy. Air may be mixed with phos- 
phorus vapor by means of stop-cock K and the characteristic 
phosphorescence will appear. 

Remarks Upon the Mitscherlich Test — Hilger and Nattermann state that 
o . 00006 gram of yellow phosphorus is the smallest quantity that can be detected 
by the Mitscherlich method. When 200 cc of water containing 0.0003 gram of 
phosphorus were distilled, there was brilliant phospLorescence for 5 minutes. 
The degree of dilution seems to have no effect upon the result, at least not within 
limits occurring in practice. Hydrogen sulphide, always present in putrefying 
animal matter, has no apparent effect upon phosphorescence. Free phosphorus 
can be detected in putrid organic matter even after the lapse of considerable time. 
Putrefactive and digestive processes appear to prevent oxidation of phosphorus. 
Dragendorff detected phosphorus in an exhumed body several weeks after death. 
Neumann found free phosphorus in a human body fourteen days and Elvers 
eight weeks after death. 

When an acid aqueous solution is distilled in the Mitscherlich apparatus, the 
flask residue always contains phosphoric (HtP04), phosphorous (HsPOi) and 
hypophosphorous (H|POi) acids and red phosphorus. Distillation of a solution 
of 0.0644 gram of phosphorus gave only 71.33 per cent, in the distillate. The 
residue contained: 

Phosphorus as phosphoric acid (H1PO4) 18.93 per cent. 

Phosphorus as phosphorous acid (H|PO|) 2.15 per cent. 

Phosphorus as hypophosphorous acid (HiPOt) 4.27 per cent. 

Phosphorus as red phosphorus 2 .oS per cent. 

28.33 

Oxidation of phosphorus may be prevented by distilling in a current of carbon 
dioxide as in the Mitscherlich-Scherer method (see page 15). 

Metabolism in Phosphorus Poisoning. — Phosphorus has a ver>' poisonous 
action upon the processes of metabolism. Present as a vapor in the blood and 
tissue fluids, it retards normal oxidative processes occurring in the animal organ- 



VOLATILE POISONS 17 

ism during metabolism. In pho^horus poisoning the usual course of chemical 
metabolism is wholly changed. Fat instead of being oxidized is deposited in the 
organs in large quantity (fatty degeneration of the liver). Different observers 
believe there is formation of fat from protein. During phosphorus poisoning the 
quantity of protein broken down is greatly increased. In hiunan metabolism 
this applies to protein in both food and tissues. Yet the needs of the organism 
are not satisfied and the conclusion is that the changes are not as complete as in 
normal protein metabolism. This increase in the breaking down of tissues in 
phosphorus poisoning recalls similar changes which take place during respiration 
in insufficient oxygen. Accompanying these abnormal processes are certain 
nitrogenous and non-nitrogenous products of metabolism which either are not 
normally formed in the organism or appear merely as intermediate steps in the 
formation of the oxidative products of metabolism. Decomposition of the protein 
molecule goes in part only as far as the amino adds. Consequently in phosphorus 
poisoning the urine almost always contains 

yen ' CHt - CH(NHs) - COOH Leucine (a-amino-isobutyl-acetic add), 
CHi^ 

>0H (i) Tyrosine (p-oxyphenyl-a-aminopropionic 

CiPK add) 

X:H, - CH(NH,) - COOH (4) 
CHt - CH(OH) - COOH Sarcolactic add (dextro-lactic add). 

In acute phosphorus poisoning the following adds can be detected in the urine 
in greatly increased quantity: 

/OH (i) 

CfHK Para-oxyphenyl-acetic add, 

^CH, - COOH (4) 

<0H (i) Para-oxy phenyl-propionic add (hydro- 

para-cumanc add). 
CH« - CH, - COOH (4) 
S - CH, - CH(NH,) - COOH 
Cystine, | , has also been detected in phosphorus 

S-CH,-CH(NH,)-COOH urine. 

In phosphorus poisoning there is a marked decrease in the urea-content of 
the urine but a dedded increase in total nitrogen. A considerable part of the 
nitrogen, that is to say, 25 per cent, or more of the total nitrogen, appears to leave 
the body as ammonia. The mine of adults usually contains from 2 to 5 per cent. 
of the total nitrogen as ammonia. The increase of ammonia may have some con- 
nection with the increase in formation of add during phosphorus poisoning. 

Peptone-like substances, the presence of which is attributed to profound dis- 
turbance of metabolism, frequently appear in the urine in phosphorus 
poisoning. Various observers believe there is no longer any doubt as to the 
appearance of genuine peptonuria. A glycosuria may also appear, or be easily 
induced by a diet rich in sugar. In accord with this observation is the fact that 
the liver of an animal poisoned by phosphorus is without the power to change 
^ucose of the blood into glycogen and store up the latter. In phosphorus 
poisoning the alkalinity of the blood rapidly diminishes owing to the increased 
formation of add. Since persons poisoned by phosphorus have icterus (jaundice), 
bile-pigment, or at least urobilin, can be readily detected in the urine. 
2 



18 



DETECTION OF POISONS 



The amounts of oiygea and carbon dioxide, which the organism respectively 
takes up and gives oS, show a marlted dinunution during phosphorus poisoning. 
Only 48 per cent, of carbon dioiide, as compared with 100 per cent, under normal 
conditions, may be eliminated. 

In brief the chief characteristics of phosphorus urine are a strong add reaction; 
presence of protein (peptone-like substances); and frequently occurrence of the 
amino acids mentioned above, as well as fat cylinders, cell detritus, free fat glo* 
bulei and blood-corpuscles. 



Fur&er Bzamination of the DistDIate 

When phosphorescence has been distinctly observed in the 
Mitscherlich apparatus, it is advisable to stop distillation and 
change the Liebig condenser to its customary position. This 
simpler method of distilling is shown in Fig. 7 and should al- 
ways be used in toxicological analysis when there is no occasioa 
to test for phosphorus. 




with liebig Condenser. 



Smce the several poisons appearing here are not equally 
volatile with steam, it is best to collect the distillate in two or 
three fractions. The first will contain most of the easily volatile 
substances like hydrocyanic acid, chloroform, ethyl alcohol, 
acetone, iodoform and nitrobenzene. The others (second and 
third) will contain substances less easily voIatUe with steam 
like carbohc acid, aniline, chloral hydrate and carbon disulphide. 
This must not be understood to mean that the first part of the 



VOLATILE POISONS 19 

distillate will be free from substances that volatilize with diffi- 
culty, and the latter part free from those that volatilize easily. 
In the main such will be the separation, but either part of the 
distillate may contain traces of substances which will appear 
in larger quantity in the other part. 

The proper procedure is to distil until 5 to lo cc. of liquid 
have been collected. Divide the distillate into several portions 
and test for hydrocyanic add, chloroform, ethyl alcohol, acetone, 
and, if necessary, also for iodoform and nitrobenzene. Use 
the second and third portions (10 to 20 cc.) to test for carbolic 
add, aniline, chloral hydrate and carbon disulphide. 

Several of these volatile substances have a characteristic 
odor, which makes it possible to recognize them with great 
certainty in the original material and especially in the distil- 
late. First, test the distillate for each individual substance 
by its most characteristic reaction. Test for hydrocyanic 
add by the Prussian blue or sulphocyanate reaction; for ethyl 
alcohol, a^cetone and acetaldehyde by Lieben's reaction; for 
carbolic add and aniline by Millon's reaction; for chloroform, 
chloral hydrate and iodoform by the phenylisocyanide reaction; 
for aniline with calcium hypochlorite solution; and finally, for 
carbon disulphide with lead acetate and potassium hydroxide 
solutions. 

When there is reason to believe that a certain substance is 
present, confirm the result by making other characteristic tests. 
It is seldom necessary to examine the distillate for all the mem- 
bers of the group. 

HYDROCYANIC ACID, HCN 

Physiolog;ical Action. — In whatever way applied, hydrocyanic acid is absorbed, 
even from the skin. So rapid is the absorption of this poison that there is 
evidence of an intoxication after a few seconds, or a few minutes at most. Part of 
the poison tHus absorbed passes from the body unchanged by way of the lungs. 
Another part, usually much less, is eliminated by the kidneys and passes into the 
urine. Sweat also is said to contain hydrocyanic acid. 

Most of the absorbed hydrocyanic acid, though variable in quantity, undergoes 
chemical change within the organism whatever be the form of its chemical com* 
bination. Hydrocyanic acid is supposed to combine with loosely bound sulphur 
of proteins and form sulphocyanic acid (HSCN) which is not nearly as toxic as 
hydrocyanic acid. (Antidote for hydroc>'anic acid.) Hydrocyanic acid after the 



20 DETECTION OP POISONS 

maimer of the Q'anohydrin reaction^ might combine chemically with carbohy- 
drates of the blood and tissues. Finally, putrefactive changes as well as ferment 
action within the cadaver might convert hydrocyanic add into ammonium for- 
mate.' The last statement may explain the disappearance of hydrocyanic add 
until only traces remain in the cadaver. Thus the possibility of making morc^ 
than an approximately quantitative determination of hydrocyanic add taken ' 
internally is preduded from the beginning. Yet there are instances where the 
poison has been found in the human cadaver after 14 days, and even after zoo and r ^ 
X 80 days. After 48 da3rs the author obtained enough hydrocyanic add in the dis- *- 
dilate from stomach and intestinal contents of a child 41/2 years old to give the 
Prussian blue test in three different portions of the distillate after 3 to 4 hours. 

Undoubtedly hydrocyanic add has a very poisonous effect upon ferments, for 
it kills certain vegetable and animal enzymes, or at least strongly retards their 
action. This acid interferes particularly with the action of that enzyme which 
causes transfer of oxygen from blood-corpusdes and thereby gives rise to oxida- 
tive processes (oxidation ferment, "respiration ferment")* Careful experiments 
in metabolism have shown that warm-blooded animals under the influence of 
hydrocyanic add take up less than the normal amount of oxygen and con- 
sequently give off less carbon dioxide, even though relatively large quantities <^ 
oxygen are administered artifidally. R. Robert (Intoxikationen) regards hydro- 
cyanic add poisoning as an internal asph>'xiation of the organs in presence of an 
excess of oxygen. The oxidative processes of the blood are checked and so little 
oxygen is used that the venous blood becomes arterial, that is to say, contains a 
large quantity of oxyhemoglobin. As a result the color of the venous blood 
is bright red. This change of venous to arterial blood seems to be permanent 
in cold-blooded but usually only transitory in warm-blooded animals. The 
appearance of lactic acid in the blood and urine is due to the disturbing influence 
of the poison upon the oxidative processes of the organism. The processes of 
normal metabolism in warm-blooded animals Anally oxidize lactic add to car- 
bon dioxide and water. Consequently the appearance of lactic add in the blood 
b ver>' transitor>' and it is not found in the urine at all. The occurrence of lactic 
add in the blood and a decrease in its alkalinity are concurrent. As a result of 
ver>' defident oxidation during hydroc>'anic add poisoning, dextrose not infr^ 
quently appears in the urine. 

The blood therefore in hydrocyanic acid poisoning is characteristically changed. 
Venous blood becomes bright red. And moreover blood which contains this add 
cannot liberate oxygen from hydrogen jwroxide, that is to say, it has lost its 
catalytic power.* Such a compound as cyano-hemoglobin appears to exbt and 



/^ 

•^-< 



-f HCN - R - C^H (R denotes any radical) 



O Hi /;0 

f H - C N - H - C; 

OH, X) - NH4 

» Hydrocyanic acid jwisons platinum black just as it does blood ferments. Put 
about 5 cc. of 3 per cent, hydmgon |>eroxide solution in each of two test-tubes. 
Add to one i or 2 dn>ps of hyilrocvanic add (about 1 per cent, solution) and to 
both a trace of platinum bUu k. Pure hydn>gcn i>eroxide at once gives off ox>-gen 
vigorously, whereas that containing hydrtKvanic acid dix?s not. 



VOLATILE POISONS 21 

its formation in the blood of a person poisoned by hydrocyanic add would seem 
probable, yet for some imknown reason the imion of this add with heemoglobin 
takes place dther not at all or only with great difficulty. 

In a chemical examination for hydrocyanic add and potas- 
sium cyanide the contents of the stomach and intestines, or- 
gans rich in blood as liver, brain and heart, the blood itself 
and sometimes the urine are most important. Examine such 
material at once for hydrocyanic add which may be recognized 
by its characteristic odor, provided putrefaction has not gone 
too far. 

Preliminary Test. — ^A spedal test (Schonbein-Pagenstecher 
reaction) for hydrocyanic add should precede distillation. 
Addify a portion of the original material in a small flask with 
tartaric add solution. Then suspend in the flask (see Fig. i) a 
strip of "guaiac-copper" paper^ without letting it touch the 
liquid. Gently warm the contents of the flask upon the water- 
bath. Neither hydrocyanic add nor potassium cyanide is 
present, unless the paper is turned blue or bluish green. But 
the only conclusion to be drawn from a positive test is that 
hydrocyanic add, or an easily decomposable cyanide, may be 
present. Further conclusions should not be drawn from a 
positive result, since other substances like ammonia, volatile 
ammonium compounds, hydrochloric add and espedally oxi- 
dizing agents like ozone, hydrogen dioxide, nitric add and 
chlorine will turn the paper blue. Consequently though very 
delicate this test cannot be accepted as condusive proof of the 
presence of hydrocyanic add. 

Mechanism of the Reaction. — Hydrocyanic acid has nothing directly to do with 
this reaction. But it forms ozone with copper sulphate and that turns the guai- 
aconic add of guaiac resin blue. Cupric cyanide (a) is an intermediate product 
which furnishes ozonized oxygen as shown in (/?): 



!j 



a) CUSO4 4- 2HCN = Cu(CN)i -f H1SO4, 
) 6Cu(CN)t + 3H,0 = 6CuCN + 6HCN -f 0|. 



* Prepare "guaiac-copper" paper by saturating strips of filter paper with 
freshly prepared, 10 per cent, alcoholic tinctiure of resin of guaiac. Dry these 
strips in air and moisten before using with very dilute aqueous copper sulphate 
solution (i : 1000). 



22 DETECTION OF POISONS 

The actual chemical examination for hydrocyanic add is made 
by adding tartaric add solution to the finely divided material 
and distilling as described (see page i8). This add volatil- 
izes easily with steam and most of it will appear in the first part 
of^the distillate. Therefore use the first 5 or 10 cc. of distillate 
for the tests. Note cautiously the odor of the distillate, which 
is characteristic, and then proceed as follows: 

!• Prussian Blue Test. — ^Add to the solution (distillate) 
a little potassium hydroxide solution; then i or 2 drops of fer- 
rous sulphate solution and i drop of ferric chloride solution. 
Shake well and warm gently. Finally addify with dilute hy- 
drochloric add. If much hydrocyanic add is present, a pre- 
dpitate of Prussian blue will appear inunediately. But if the 
quantity is small, the solution will have merdy a blue or bluish 
green color. After a long time (10 to 12 hours) a flocculent 
predpitate of Prussian blue will settle to the bottom of the test- 
tube. The limit of delicacy is i 15,000,000.^ 

Mechanism of the Reaction.' — Hydrocyanic acid and potassium hydroxide 

form potassium cyanide which with ferrous sulphate produces ferrous cyanide 

(a). The latter combines with more potassium cyanide, forming potassiiun 

ferrocyanide (/3) which with ferric chloride precipitates Prussian blue (7), the 

ferric salt of hydroferrocyanic acid (H4Fe(CN)e). 

(a) FeS04 + 2KCN = Fe(CN), + KjSO*, 

09) Fe(CN), + 4KCN = K4Fe(CN)6, 

(7) 3K4Fe(CN)6 + 4FeCl, = Fe4[Fe(CN)6]i + 12KCI. 

Prussian blue will not appear in presence of alkalies, since they decompose it as 

follows: 

Fe4[Fe(CN)J, -h 12KOH = 3K4Fe(CN)e -h 4Fe(0H),. 

Consequently test the final mixture with blue litmus paper to make sure it is acid. 

2. Stdphocyanate Test. — Add to a portion of the distillate 
3 or 4 drops of potassium hydroxide solution and then a little 

* Link and M5ckel, Zeitschrift fiir analytische Chemie 17, 455 (1878). 

* The reaction may be explained by saying that all the ferrous salt in presence of 

much potassium hydroxide is precipitated as ferrous hydroxide (a). The latter 

with potassium cyanide then forms ferrous cyanide (0) and finally potassiiun 

ferrocyanide (7) : 

(a) FeS04 -h 2KOH = Fe(OH), + K,S04, 
09) Fe(OH), + 2KCN = Fe(CN), + 2KOH, 
(7) Fe(CN), + 4KCN - K4Fe(CN)e. 

With ferric chloride potassium ferrocyanide in presence of hydrochloric add 
finally gives Prussian blue (see above). 



VOLATILE POISONS 23 

yellow ammoniiun sulphide solution. Evaporate to dryness 
upon the water-bath. Dissolve the residue in a little water, 
and acidify with dilute hydrochloric add. Filter through 
double paper to remove sulphur, and add to the filtrate 2 or 3 
drops of ferric chloride solution. If the distillate contained 
hydrocyanic add, a reddish to blood-red color will appear. 
This is due to ferric sulphocyanate. The limit of delicacy is 
1 : 4,000,000. 

Mechanism of the Reaction. — Hydrocyanic add and potassium hydroxide 

form potassium cyanide which takes sulphur from yellow ammonium sulphide 

and becomes potassium sulphocyanate (a). The latter with ferric chloride forms 

ferric sulphocyanate (/?): 

(a) KCN + (H4N)jS, - KSCN + (H4N),S,-i, 
0?) 3KSCN + FeCl, - Fe(SCN), + 3KCI. 

3. Vortmaxm's^ Nitroprusside Test — Add to a portion of the 
distillate a few drops of potassium nitrite solution; then 2 to 4 
drops of ferric chloride solution and enough dilute sulphuric 
acid to give a bright yellow color. Heat to boiling, add suf- 
ficient ammonium hydroxide solution to remove excess of iron 
and filter. Add to the filtrate i or 2 drops of very dilute am- 
monium sulphide solution. If the solution contained hydro- 
cyanic acid, a violet color will appear and pass through blue, 
green and yellow. The limit of delicacy is i 131 2,000. 

Note. — This test is the reverse of the nitroprusside test for hydrogen sulphide 
and is due to conversion of hydrocyanic add to potassium nitroprusside, K|Fe- 
(NO)(CN)s, which causes the color changes when ammonium sulphide is added. 
Very small quantities of hydrocyanic acid give a bluish green to greenish yellow 
color. 

4* Silver Nitrate Test. — Acidify a portion of distillate with 
dilute nitric acid, and add silver nitrate solution in excess. 
If hydrocyanic add is present, a white, curdy precipitate of 
silver cyanide (AgCN) will appear. Excess of ammonium 
hydroxide solution will readily dissolve this precipitate. The 
limit of delicacy is i : 250,000. 

When a dilute aqueous solution of hydrochloric add is dis- 
tilled, the add does not pass into the distillate. The pre- 
dpitate, therefore, caused by silver nitrate solution cannot 

^ Monatshefte fttr Chemie 7, 416 (1886). 



24 DETECTION OF POISONS 

• 

possibly be sUver chloride, because at first nothing but pure 
water distils from a i per cent, or weaker solution of hydro- 
chloric acid. To remove hydrochloric acid, if present, re- 
distil the first distillate over borax. This will retain hydro- 
chloric but not hydrocyanic acid. It is advisable to collect 
upon a filter, wash and dry the precipitate caused by silver 
nitrate solution. If silver cyanide is heated in a bulb-tube, it 
will form metallic silver and cyanogen gas. The latter may be 
recognized by its charactertisic odor.^ The reaction is: 

2AgCN » 2Ag + (CN)i. 

5. Picric Acid Test — Make alkaline a portion of distil- 
late with potassium hydroxide solution and heat gently (50 to 
60°) with a few drops of picric acid solution. If hydrocyanic 
acid is present, the solution will become blood-red. This is due 
to formation of potassium isopurpurate. 

Note. — ^This test is not as delicate, nor as characteristic of hydrocy anic add 
as the other tests. If hydrogen sulphide is present, as it frequently is in distillates 
from animal matter, picric acid solution will produce a red color owing to forma- 
tion of picraminic acid. 

6. Weehuizen^s^ Test — ^Add a few drops of phenolphthalin 
dissolved in dilute sodium hydroxide solution and then a little 
copper sulphate solution (i : 2000) to a portion of the distillate. 
Even in a dilution of i : 500,000 hydrocyanic acid will produce 
a red color due to oxidation of phenolphthalin to phenolphthalein. 
Such oxidizing agents as hydrogen peroxide, nitric add and 
ferric chloride do not give this test. Paper first moistened with 
alkaline phenolphthalin solution and then with very dilute copper 
sulphate solution may be used. These phenolphthalin-copper 
sulphate papers turn red even in air containing hydrocyanic acid. 

Under the conditions of the test phenolphthalin is oxidized to phenolphthalein: 



. H 
+ 

-.. H 



/CeH^—OH /CeHc-OH 

— CfC«H^--OH - C^^CeHc-OH + H,0 
\CeH4 I ^CeH4 

O— OC 0— CO 



Phenolphthalin Phenolphthalein 

^ Owing to the very poisonous character of cyanogen gas, it is safer to ignite the 
gas at the mouth of the bulb-tube. Cyanogen gas bums with a purple flame. 
Tr. 

* Pharmaceutisch Weekblad 43, 271; and Pharmaceutische Zentralhalle 46, 

256 (1905)- 



I VOLATILE POISONS 25 

I On the otber hand the phthalein heated with an alkaline hydroxide and xincdust 
I ii reduced to the phtbalis. 

I QuantitatiTe EEtimatioii of Hydrocyanic Acid 

' To determine hydrocyanic acid quantitatively, acidify ft weighed portion of 
material with dilute sulphuric or tartaric acid and distil. Detemiine the 
quantity of hydrocyanic acid in the distillate either gf a vi metrically or volu- 
metrically. If the former method is used, collect the precipitate of ^Iver cyanide 
upon a weighed filter, wash and dry at ioo° to constant weight; or ignite the 
precipitate in a weighed porcelain crucible, and determine the quantity of me- 
tallic silver obtained. If hydrochloric add is present in the distillate, rediatil 
once over borax. The distillate will then be free from hydrcM:hIoric acid. 

I Detection of Hydrocyanic Acid in Presence of Potaasium Feirocyanide 

r When material contains aon-pobonous potassium ferrocyanide, hydrocyanic 
ftdd will appear in the distillate from a solution acidified with tartaric add. In 
V) experiment, where r per cent, potassium ferrocyanide solution was distilled 
with 0.03 gram of tartaric acid, the distillate contained considerable hydrocyanic 
add. Carbon dioxide, passed into hot, aqueous potassium ferrocyanide solution, 
will liberate hydrocyanic acid even at water-bath temperature (73°). To test for 
potassium ferrocyanide beforehand, shake some of the original material with 
water and filter. Test the filtrate with ferric chloride solution and dilute hydro- 
chloric add. n there is a predpitate of Prus^au blue, potassium ferrocyanide is 
present. To detect free hydrocyanic add, potassium or sodium cyanide' with 
certainty, in presence of potassium ferrocyanide, add to the material add sodium 
carbonate in not too small quantity and distil. Even long dlslillation over free 
^me by this method will liberate hydrocyanic add only from simple cyanides and 

r not from potassium ferrocyanide. 

r Detection of Mercuric Cyanide 

When an aqueous solution of mercuric cyanide, which is exceedingly poisonous, 
is distilled with tartaric add, the distillate will contain hydrocyanic add only 
when a large quantity of mercuric cyanide is present. Distillation of 100 cc. of 
I per cent, aqueous mercuric cyanide solution yields a distillate which gives the 
Prussian blue test distinctly. But, if the quantity of mercuric cyanide is less and 
the solution very dilute {for example, 100 cc. oEo.ot per cent, solution), there will 
not be a trace of hydrocyanic add in the distillate, even though the solution is 
Otioniily addified with tartaric add. If. however, a few cc. of freshly prepared 
hydrogen sulphide water are added and distillation is resumed, mercuric cyanide 
will be completely decomposed and the distillate will contain bydrocyaaic add. 

Detection of Mercuric Cyanide in Presence of Potassium Ferrocyanide 

The method of detecting hydrocyanic acid from simple cyanides, in presence of 
potassium ferrocyanide, is not applicable to mercuric cyanide. Long distillation, 
even from saturated add sodium carbonate solution, gives no trace of hydrocyanic 
•cid. But dbtitlatioa in presence of not too little add sodium carbonate, alter 
addition of a few cc. of freshly prepared, saturated hydrogen sulphide solution, 
rcuric cyanide is an exception. 



26 DETECTION OF POISONS 

liberates hydrocyanic acid from mercuric cyanide but not from potassium 
ferrocyanide. It is possible to detect hydrocyanic acid by this method, when 
very little mercuric cyanide is mixed with considerable potassium ferrocyanide. 
For example, o.oi gram of mercuric cyanide in loo cc. of lo per cent, potassium 
ferrocyanide solution can be detected. If potassium ferrocyanide is distilled 
directly with hydrogen sulphide without addition of acid sodium carbonate, 
the distillate will contain considerable hydrocyanic acid. 

CARBOLIC ACm 

Action and Fate of Carbolic Add in tiie Animal Body 

Concentrated carbolic acid coagulates and destroys the constituents of the 

human body, especially proteins and protoplasmic structures. It has therefore a 

very strong caustic action. But its action is not merely local, for after absorption 

OH it shows an affinity particularly for the central nervous S3rstem, 

1 brain and spinal cord. The first indications of this in animals are 

jr\ strong stimulation, increased irritability as in the case of strych- 

HC CH nine and paral>'sis. In man the period of stimulation is very 

I slow in appearing. In chronic poisoning, after repeated small 

"^ y doses of carbolic acid, degeneration of the kidneys and liver is a re- 

{^ suit of absorption. The human organism absorbs carbolic add 

H very rapidly. Absorption from the skin, the gastro-intestinal 

tract, abrasions and the respiratory organs takes place readily. In the himian 

organism the poison is converted by conjugation with sulphuric acid into phenyl- 

sulphuric acid: 

HO-SOi-OH -f IIO-CeH* - H0-S0rOC«H* + H,0. 

When the quantity of carbolic acid is ver>' large, it is also converted into phenyl- 
glycuronic acid by conjugation with glycuronic add, HOOC-(CH.OH)4CHO. 
Considerable cjirl)i)lii' uciil is oxidizeil within the body to dihydrox>*-bcnaenes, 
namely pyrtnatechol (C6H4((^H)i(i»a)) and hydroquind (C»H4(OE0i(i,4)). 
These enter into Hynthe»is with sulphuric acid and appear in urine as ethereal 
salts of sulphuric add, 'l*he dark o>lor of "carbolic urine" is largdy due to 
further oxidation of hydnxiuinol, whereby colored products (quinone ?) are 
formeii. In carbolir add poisoning, urine often has a pronounced daric color 
(greenish to blade). Vrlne in other cases is amber->'ellow at first, but standing in 
air gives it a deeper ittlor. When carlu^lic acid poisoning is suspected, the urine 
should be examined rhemiially. **CarlH\lic urine** differs from normal human 
urine in being nearly free frt>n) sulphuric acid,^ the so-called "preformed sul- 
phuric add." (\mHe()nently barium ihloride solution^ in presence of excess of 
acetic add, gives only a sliKht precipitate of barium sulphate or none at all. 
Miter when there U a predpitate and warm the dear filtrate with a few cc of 
concentrated hvdroi hlortr aihl. An abundant pTrd}^tate of barium sulphate 
will usually appear. The mineral adtl de^xun^H'^ses phenyUsulphoric add into 
phenol and Mulphuiii adil whirh is then prtnii^itated. Normal human urine 

^ This is iulphuili a^ Id pienent in mine as sulphates* It is also termed "pre- 
formeil sulphuili' ac Itl," \\\ whldi I* meant that it enters the body as such. In 
this respect it dilYeis liom '*0thrieal/' or *\0ivjucate** sulphuric adds, whidi 
retult frtmt synlhrHCM within the biuly, 



VOLATILE POISONS 27 

contaiiis considerably more ''sulphate sulphuric acid" (A — sulphuric acid) than 
" ethereal sulphuric add " (B — sulphuric add) . The average proportion between 
the two being: A — S04:B — S04=io:i. Barium chloride solution, added to 
normal urine in presence of acetic acid, produces a heavy predpitate of barium 
sulphate. 

Distribution of Carbolic Add in the Human Body After Poisoning 

C. Bischofif^ examined organs, removed from a man who died 15 minutes after 
taking 15 cc. of liquid carbolic acid, and foimd the poison distributed as stated in 
the table below. The organs in this case were perfectly fresh. Only a small por- 
tion of the stomach was received. 

Weight Organ Phenol 

242 grams Contents of stomach and intestine 0.171 gram 

113 grams Blood 0.028 gram 

1480 grams Liver 0.637 gram 

322 grams Kidney 0.201 gram 

1445 grams Brain 0.314 gram 

Bischoff distilled with steam until the distillate gave no further predpitate with 
bromine water. The results show how rapidly carbolic acid is absorbed, and how 
soon it is distributed throughout the body. 

£. Baumann' has published certain facts relating to the quantity of carbolic 
add formed during putrefaction of protein substances. Baumann states that he 
obtained from 100 grams of fresh pancreas and 100 grams of moist fibrin, mixed 
with 250 cc. of water, after 6 days of putrefaction 0.073 to 0.078 gram of tri- 
bromophenol, corresponding to 0.0208 to 0.022 gram of phenol. 

Urine gives a distinct test for carbolic add 15 minutes after the poison has 
been taken by the mouth, or hypodermically. This shows how rapidly carbolic 
add is absorbed. Most of the carbolic acid absorbed is eliminated in 4 or 5 
hours. Schaffer* found the quantity of conjugate sulphuric acid in urine to 
increase in exact proportion to the quantity of carbolic add taken. 

Tests for the Detection of Carbolic Acid 

Carbolic add distils quite easily with steam. Yet, to remove 
the last traces, long-continued distillation is necessary. If 
fractional distillation is made, when carbolic acid is present, 
this substance will appear in the first and second fractions and 
even in the third. Usually carbolic acid can be recognized by 
its'peculiar odor. When much carbolic acid is present, the 
distillate is milky. Colorless or reddish globules may be seen 

* Berichte der Deutschen chemischen Gesellschaft 16, 1337 (1883). 

* Berichte der Deutschen chemischen Gesellschaft 10, 685 (1877) and Zeitschrift 
ffir physiologische Chemie i, 61 (1877-78). 

'Journal fur praktische Chemie, Neue Folge 18, 282 (1878). 



28 DETECTION OF POISONS 

floating in the liquid. Excess of potassium or sodium hydroxide 
solution will dissolve carbolic acid and render the distillate 
perfectly clear. Pure, anhydrous carbolic acid melts at 40 
to 42° and distils at 178 to 182°. 

Decomposition of protein substances produces phenol and 
especially para-cresol in small quantity. Traces of phenols 
can almost always be detected in distillates from animal matter 
in an advanced stage of decomposition. Millon's reagent, 
and usually bromine water, will give positive tests with such 
distillates. 

I. Millon's Test. — Millon's reagent,^ heated with a solution 
containing only a trace of carbolic add, produces a red color. 
An aqueous solution containing only 20 mg. of carbolic acid, 
diluted 1 : 100,000, will give a distinct red color. If the phenol 
solution is not very dilute, the color will appear even in the cold. 
Though a very delicate test, it is not characteristic of carbolic 
acid, because several other aromatic compounds behave simi- 
larly. This is true of derivatives of mon-add phenols like the 

three cresols, salicylic add,' para- 
hydroxy-benzoic add, para-hydroxy- 
phenyl-acetic add, para-hydroxy- 
phenyl-propionic add (hydro-para- 
cumaric add') and tyrosine. Aniline 
heated with Millon's reagent also 
gives a dark red color. 

2. Bromine Water Test. — Excess 
of bromine water produces a yellow- 
ish white, crystalline predpitate, 

Fig 8.-Tribromophenol ^^^^ ^j^ ^^^^ CarboUc add 

Crystals. From a dilution . ; 

of 1 : 20 000. solutions. It IS a very dehcate test 

for carbolic add. Phenol diluted 
1 : 50,000 yields, after some time, a precipitate made up in 
part of well-formed crystals (Fig. 8). 

^ For the preparation of this reagent see page 314. 

* Traces of salicylic acid volatilize with steam, at least in such quantity that it 
can be detected with Millon*s reagent. 

> Para-hydroxy-phenyl-acetic acid and hydro-para-cumaric acid are formed in 
the putrefaction of proteins but are not volatUe with steam. 




VOLATILE POISONS 29 

If there is a sufficient excess of bromine water to give the supernatant liquid a 
brownish red color, the precipitate consists only of tribromophenyl hypobromite, 
CsHiBriO. R. Benedikt^ regards this compound as a brom-phenoxy-tribromo- 
benzene with the structure 



i 



OBr , whereas Thiele and Eichwede* have ascribed to it the structure 

O 

/x s 

BrC CBr /\ 

II I BrC CBr 

HC CH II 11 

\^ HC CH 

^ V 

Br C 

Br, 

This reaction takes place so easily that carbolic add may even be determined 

quantitatively as this tetrabromo-derivative (see page 31). It melts at 132- 

^H 124® with evolution of bromine and crystallizes as lemon-yellow 

X leaflets from alcohol-free chloroform or ligroin. Heated with 

^\^ alcohol, acetone, xylene, or aqueous sulphurous add, this com- 

BrCf^aCBr pound loses bromine and changes at once to 2,4,6-tribromo- 

I 1 phenol, melting at 93-94®. Salicylic alcohol (saligenin), sali- 



\ 



y cylic aldehyde, salicylic add and para-hydroxy-benzoic add 

(5^ are converted quantitativdy by an excess of saturated bromine 



Br water even in the cold into tribromo-phenyl hypobromite. 

3^ Ferric Chloride Test. — Very dUute ferric chloride solution, 
added drop by drop, imparts a blue-violet color to aqueous 
carbolic add solutions. Addition of dilute hydrochloric or 
sulphuric acid changes this color to yellow. This test is not as 
delicate as i and 2. It is entirely negative in presence of min- 
eral adds. The limit of delicacy is about i : 1000. 

4* Hypochlorite Test. — Add a few cc. of ammonium hydrox- 
ide solution to a dilute, aqueous carbolic add solution, and then 
2 or 3 drops of freshly prepared caldum or sodiimi hypochlorite 
solution. Gentle warming will produce a blue color. Very 
dilute carbolic acid solutions after some time give only a green 
to blue-green color. F. A. Fluckiger' allows bromine vapor 
to come into contact with the phenol solution which has been 
mixed with a little ammonium hydroxide solution in a porcelain 
dish. 

^ Annalen der Chemie und Pharmade 199, 127 (1879). 
*Berichte der Deutschen chemischen Gesellchaft 33,637 (1900). 
' Pharmaceutische Chemie, page 287 (1879). 



30 DETECTION OF POISONS 

$• Nitrite Test — Mix a carbolic add solution with a dilute 
alcoholic solution of ethyl nitrite, C2H6-0-N=0,^ or iso- 
amylnitrite, C6Hii-0-N = 0,^ and add concentrated sulphuric 
acid from a pipette so that it forms a distinct under-layer. A 
red zone will appear at the contact surface of the two liquids. 
This is a very delicate test. 

This test may also be made by adding the liquid under ex- 
amination as an upper layer upon concentrated sulphuric acid 
containing a trace of red fuming nitric acid. 

6. H. Melzer's Benzaldehyde Test' — ^Add 2 cc. of concen- 
trated sulphuric acid to i cc. of the solution (distillate) to be 
tested for carbolic acid, then i or 2 drops of benzaldehyde and 
heat. The mixture, at first yellowish brown, will become dark 
red. At the same time a red resinous substance will appear, 
unless the solution is too dilute. When cold add 10 cc. of water 
and enough potassium hydroxide solution to give a distinct 
alkaline reaction. If carbolic acid is present, a violet-blue 
color will appear. To obtain this coloring-matter, acidify the 
solution, extract with ether and evaporate the solvent. Alka- 
lies, added to alcoholic solutions of the coloring^matter, produce 
a blue color which acids discharge. This is a very delicate 
test. One cubic centimeter of 0.05 per cent, carbolic acid solu- 
tion (s^ 0.0005 gram of carbolic acid), will still give the blue 
color very distinctly. 

Note. — In absence of phenol concentrated sulphuric acid product a dark 
brown color with benzaldehyde. According to A. Russanow^ the first condensa- 
tion product between phenol and benzaldehyde in presence of concentrated 
sulphuric acid is para-dihydroxy-triphenyl-methane which crystallizes in yellow- 
ish needles: 

C.H6V HC«H4-0H CeHs CeH4-0H 

>C=iO+ =H,0+ >C< (1,4). 

Be&caldehyde Phenol P-Dihydrozy-triphenyl>methane 

Akalies dissolve the pure crystals without color but, if these solutions are 
exposed to air, oxidation takes place and a red or red- violet color appears. Prob- 

1 The officinal preparation is called "Spiritus Aetheris Nitrosi." 

« Amyliiun nitrosum of pharmacists. 

« Zeitschrift fur analytische Chemie 37, 345 (1898). 

* Berichte der Deutschen chemischen Geselbchaft 22,1943 (1889). 



VOLATILE POISONS 31 

ably benzaurine, dihydroxy-triphenyl-carbinol, is first formed. This compound 
is a brick-red crystalline powder soluble in alkalies with a violet color. 

QUANTITATIVE METHODS OF ESTIMATING PHENOL 
z. Gravimetric Estimation as Tribromophenol 

The principle of this method is based on the complete pre- 
cipitation of phenol from aqueous solution as tribromophenyl 
hypobromite by an excess of saturated bromine water (see 
Test 2) . The precipitate is practically insoluble in cold bromine 
water and the results are very satisfactory.^ 

Procedure. — Place the aqueous phenol solution in a large 
glass-stoppered flask. Add gradually, while shaking, saturated 
bromine water until the supernatant liquid has a red-brown 
color and bromine vapor is visible above the solution. Let 
stand 2-4 hours and shake frequently. Then collect the pre- 
cipitate in a weighed Gooch crucible and dry in a vacuum des- 
iccator over sulphuric add to constant weight. On the basis of 
the following proportion calculate the weight of phenol corre- 
sponding to the weight of the precipitate: 

CeH|Br40 : CJIg-OH = Wt. of Ppt. found :x 
409.86 94*05 

Since the ratio ^~~n^ = 0.2295, the weight of phenol may be 
found by multiplying the weight of the precipitate by 0.2295. 

2. Beckurts-Koppeschaar> Volumetric Method 

Dilute sulphuric add liberates hydrobromic add from potas» 
sium bromide (a) and bromic add from potassium bromate (/S). 
These two acids react according to (7) : 

(a) KBr + HjSO* = KHSO4 -f HBr, 
03) KBrO, + H,S04 = KHSO4 + HBrO,, 
(7) sHBr + HBrO, = sBr, + 3H1O. 

* The following results were obtained by F. Beuttel: 

Phenol taken CtHsBr40 Phenol found Per cent, found 

X. 0.103 grm. 0.4538 grm. 0.0997 grm. 96.2 

2. 0.2072 grm. 0.8806 grm. 0.2014 grm. . 98.6 

3. 0.2072 grm. 0.8708 grm. 0.2006 grm. 98.6 

* Archiv der Pharmazie 24, 570 (1886). 



32 DETECTION OF POISONS 

Therefore addition of dilute sulphuric add to a mixture of 
potassium bromide and bromate solutions . liberates bromine 
which will convert phenol into a mixture of tribromophenol and 
tribromophenyl hypobromite. The excess of free bromine 
and also the loosely bound bromine atom of tribromophenyl 
hypobromite will displace iodine from potassium iodide and 
finally all the phenol will be present as tribronHphenol: 

CeHsBnOBr + aKI - CeHsBfiOK + KBr +Ii 

One molecule of phenol requires 6 atoms of bromine, as shown 
by the equation: 

sKBr + KBrOt + 6H,S04 + CeH,OH - CaHtBrsOH + sHBr + 6KHSO4 + 

3H,0. 

The following standard solutions are required: 

1. o.oi n-potassium bromide solution, containing 

5Q5-6 , ^^ . 

— -Lzv_ _ 5.956 grams KBr m 1000 cc. 

2. o.oi n-potassium bromate solution, containing 

167.17 , ^_ ^ . 

grams = = 1.67 17 grams KBrOa m 1000 cc. 

3. 0.1 n-sodium thiosulphate solution, containing o.i 
Na2S203.5H20 grams = 24.83 grams in 1000 cc. 

4. Potassium iodide solution, containing 125 grams of KI 
in 1000 cc. 

Procedure. — Put about 25 cc. of aqueous phenol solution 
(distillate) into a flask having a tight glass stopper. Add 50 
cc. each, of o.oi n-potassium bromide and o.oi n-potassi\mi bro- 
mate solutions, then 5 cc. of pure concentrated sulphuric acid 
and shake vigorously for several minutes. The gradually 
increasing opalescence of the solution becomes more and more 
marked, as tribromophenol and tribromophenyl hypobromite 
are precipitated. The yellow color which soon appears shows 
excess of bromine. Open the flask in 15 minutes, add 10 cc. 
of potassium iodide solution, shake and titrate free iodine in 
5 minutes with o.i n-sodium thiosulphate solution. 



grams 

^ 100 



VOLATILE POISONS 33 

6 gram-atoms Br 6 X 79*96 

CalcolatiQii. = = 4.7976 grams of bromine are 

100 100 

set free from a mixture of 1000 cc. of o.oi n-potassium bromide solution and 1000 
cc of 0.01 n-potassium bromate solution. A mixture therefore of 50 cc. of each 
of the two solutions will give 0.3399 gram of bromine. This quantity of bro- 
mine can convert 0.04704 gram of phenol into tribromophenol: 

6Br : C,H,OH 
479.76 9405 - 0.2399 : X (x » 0.04704) 

z cc. of o.i n-sodlRn thiosulphate solution corresponds to 0.012697 gram of 

iodine and this quantity of iodine to 0.007996 gram of bromine. But 0.007996 

gram of bromine will convert 0.00157 gram of phenol into tribromophenol: 

6Br : CeH»OH 

479.76 94.05 — 0.007996 : X (x — 0.00157) 

Consequently, for each cc. of o.i n-sodiimi thiosulphate solution used, subtract 
0.00157 from 0.04704 gram of phenol. This determines the quantity of car- 
bolic add in the 25 cc. of distillate taken. 

3. Messiiiger-Vortmaim^ Volumetric Method 

Excess of iodine (8 atoms of iodine to i molecule of pheno 
dissolved in 4 molecules of potassium hydroxide) , added to an 
alkaline phenol solution at 50-60°, will produce a dark red, non- 
crystalline precipitate. One molecule of phenol requires 6 
atoms of iodine: 

1. CeHftOH + 3I1 = C»HJ,OH + 3HI. 

2. 3HI + 3KOH = 3H,0 + 3KI. 

This red precipitate dissolves in hot potassiimi hydroxide 
solution with a red-brown color and appears as white 2, 4, 6-tri- 
iodophenol, melting at 154-156°, on addition of an excess of 
dilute sulphuric acid. Messinger and Vortmann regard the 
red compound as di-iodophenyl hypoiodite (C6H8I2OI) which 
potassium hydroxide converts into the more stable isomeric tri- 
iodophenol : 

01 OH 

c c 

j^\ is converted by ^\ 

IC CI potassium hy- IC CI 

I II drozide into | || 

HC CH HC CH 

Y Y 

H I 

Red di-iodo- White 2, 4, 6-tri- 

phenyl hypoiodite iodophenol 

^ Berichte der Deutschen chemischen Gesellschaft 22, 2312 (1889); and 23, 
3753 (1890). See also Kossler and Penny, 2^tschrift fttr physiologische Chemie 
17, 117 (1892). 
3 



34 DETECTION OF POISONS 

Procedure.^ — The reaction between the alkaline phenol solu- 
tion and iodine is rather slow in the cold but is hastened at 50 
to 60°. 

» 

Place a measured volume of aqueous phenol solution (5 to 
10 cc.) in a small flask and add a measured volume of o.i 
n-potassium hydroxide solution until the mixture is strongly 
alkaline. Warm gently by dipping the flask in water at 60° 
and add 10-15 cc. more of o.i n-iodine solution than the 
volume of o. i n-potassium hydroxide solution used, or until the 
excess of iodine produces a strong yellow color. Agitation will 
cause a deep red precipitate to appear. Cool the solution, 
acidify with dilute sulphuric add and dilute to a definite volume 
(250 to 500 cc.) . Filter an aliquot portion (100 cc.) rapidly and 
determine excess of iodine with 0.1 n-sodium thiosulphate solu- 
tion. 

Calculation. — Each molecule of phenol requires 6 atoms of 

iodine. Therefore i atom of iodine = j = ' = 

o o 

15.675 phenol. 1000 cc. of 0.1 n-iodine solution, containing 
O.I gram-atom of iodine, correspond therefore to 1.5675 grams 
of phenol. 

Note. — This method will not give satisfactory results, unless 
at least 3 molecules of sodium or potassium hydroxide are taken 
for I molecule of phenol. 

EstimatiQn of Phenol in Urine 

In determining carbolic add in urine, the regular occurrence of phenols must 
not be overlooked. After a mixed diet, the quantity of normal human urine 
passed in 24 hours will yield approximately 0.03 gram of phenols (phenol and 
more especially para-cresol). 

In certain diseases where there is excessive bacterial decomposition within the 
organism, in the intestines for example, urine contains more of these phenols and, 
consequently, more conjugate sulphuric adds. Even external application of 
carbolic add, for instance the use of carbolic add water as a lotion, is suffident to 
increase the quantity of phenyl-sulphuric add in urine. 

Detection of Carbolic Add in Presence of Aniline 

Aniline dosely resembles carbolic add in behavior toward Millon's reagent and 
bromine water. But the two substances can be easily separated. Add potassium 

* Use 0,5 to I per cent, carbolic add solution for laboratory experiments. 



CHLOROFORM 



VOLATILE POISONS 35 

' liydtoxide solution in large excess and distil. The distillate will contain aniline 
alone. Or make the solution strongly add with dilute sulphuric add, 
and eitract with ether which will dissolve only tacbolic acid. Evaporate 
the etber extract at a moderate temperature and examine the residue. 

^^1 Behavior in the Hunun Orgamsm. — When inhaled chloroform first passes from 
^^■Sw air into the btood-pksma which then transmits it to the red blood-corpusdes 
^^» jj where it may accumulate in relatively large quantity. Air passed 

^^K^ I through blood will remove chloroform completely. Pohl (see 

^ — C — CI Kobert's " Intoxibationcn ") states that blood may contain 0.63 
^^B L per cent, of chloroform, three-fourths of which will be in the red 

^^B blood-corpuscles. At the height of a harmless narcosis the blood 

^^pDOntained only 0.035 P^^ cent, of chloroform. Absorption of cliloroform is rapid 
from all parts of the body. The stimulative action of chloroform on the mucous 
membranes of the respiratory passages explains such disturbances as coughing, 
secretion of saliva and reflex slowing of respiration and heart-beat, occurring at 
the beginning of narcosis. Dilatation of the biood-vessels of organs living after 
death is due to paralysis caused by even small doses of chloroform, A drop in 
blood-pressure accompanies paralysis of the brain and the heart's action is feebler 
and slower. Several researches regarding the effect of inhaled chloroform upon 
human and animal metabolism have shown an increase in the quantity of nitrogen 
in the urine alter prolonged narcosis because moce protein ia decomposed. The 
amount of neutral sulphur and chlorine in the urine also increases. The increase 
of the latter is due in part at least to the conversion ol chloroform into chloride. 
The addity of the urine b also much higher. A final characteristic of chloroform 
urine is the high content of reducing substances. The increased protein decom- 
position in chloroform narcosis aficcts both reserve protein and that of the tissues. 
This may explain degeneration in red blood-corpusdes, gbndular organs, the 
heart, etc., after frequent narcoses or one of long duration. 

The temporary or permanent paraly^s of isolated animal or vegetable cells, 
such as leucocytes, dilated cells, yeast cells, algEG and spores. Is evidence of the 
antiseptic action of chloroform when present in properconcentrationinair orina 
liquid. This enplains the use of i per cent, aqueous chloroform solution as an 
antiseptic. Added to urine it acts as a preservative. Therefore it may be used 
in the study of the action of enzymes but not of bacteria, though all micro-organ- 
isms are not paralyzed or killed by chloroform water. 

Pohl and Hans Meyer have studied the distribution of chloroform in the body 
and found that the red blood-corpuscles and the brain are most likely to show this 
person. After chloroform has been inhaled, some wil! appear in the gastric juice 
but at most only traces in the urine. In but two out of 15 cases of chloroform 
narcosis was this poison found in the urine and then only in traces. 

Eobert states that as a rule it is the exception to find chloroform itself in the 
cadaver, because part of the poison is converted into chlorida in the human organ- 
ism and part is quickly exhaled during respiration. Usually it is possible to detect 
dllorofonn in the breath of patients even 34 hours after narcosis. BUdingei 
tatei that the mucus of the respiratory passages retains chloroform. 



36 DETECTION OF POISONS 

Tests for the DetectiQn of Chlorofonn 

Chloroform distils easily with steam and appears in the first 
fraction in largest quantity. When much chloroform is present, 
it will separate from the distillate as heavy, colorless globules, 
whereas a small quantity will remain in solution. This solution 
usually has the characteristic odor and sweetish taste of chloro- 
form. The following tests should be applied to the first frac- 
tion. 

1. Phenylisocyaside Test. — Add i or 2 drops of aniline to the 
chloroform solution (distillate), and then a few cc. of aqueous, 
or alcoholic potassium hydroxide solution. Gentle heat will 
produce phenylisocyanide (CeHsNC). The penetrating and 
very repulsive odor of this compound is easily recognized. 

CHCli + CeH,.NH, + 3KOH = CeH|.NC + 3KCI + 3H,0. 

A. W. Hofmann states that this test will show with certainty 
I part of chloroform in 5000 to 6000 parts of alcohol. 

Note. — Chloral, chloral hydrate, bromoform, iodoform and tetrachloro- 
methane also give this test. 

The fact that aniline boiled with potassiiun hydroxide solution gives a peculiar, 
faintly ammoniacal odor, even when chloroform is absent, must not be over- 
looked. There is small chance, however, of confusing this odor with the repulsive 
smell of phenylisocyanide. In doubtful cases warm some water, containing a 
drop of aniline and a trace of chloroform, with potassium hydroxide solution and 
compare the odor with that in question. 

2. Schwarz's Resorcinol Test.^ — Dissolve about o.i gram of 
resordnol ( C6H4 \pvTT / n ) in 2 cc. of water, add a few drops of 

sodium hydroxide solution and finally the liquid containing 
chloroform. This mixture heated to boiling will develop even 
in very dilute solution a yellowish red color attended by a 
beautiful yellowish green fluorescence. 

Chloral, bromal bromoform and idoform also give this test. 

3. Lustgarten's^ Naphtho] Test. — Dissolve a few centigrams 
of a- or /3-naphthol in i or 2 cc. of 33 per cent, aqueous potas- 
sium hydroxide solution. Warm to 50® and add the solution to 

* Zeitschrift ftlr analytischc Chemie, 27, 668. 
" Monatshefte fUr Chemie, 3, 715 (1882). 



VOLATILE POISONS 37 

be tested. Chloroform will produce an evanescent blue color 
which in contact with air will change to green and then to 
brown. This color is less stable when /3-naphthol is used. 
Acidification of the blue solution will precipitate naphthol col- 
ored by a red dye stuff. This precipitate is usually brick-red. 

Chloral, bromal bromofonn and idoform also give this test. 

4. Cyanide Test. — Seal the liquid to be tested for chloroform 
in a glass tube (pressure-tube^) with a little solid ammonium 
chloride and alcoholic potassium hydroxide solution. Heat 
for several hours in a boiling water-bath. Cool the tube, re- 
move the solution and test for hydrocyanic acid by the Prussian 
blue reaction. A positive test means that the distillate con- 
tained chloroform. The following reactions take place: 

(a) CHCl, + H,N + 3KOH = HCN + 3KCI + 3H,0, 
03) HCN + KOH = KCN + H,0. 

5. Reduction Tests, (a) With Fehling's Solution. — Warm 
the liquid containing chloroform with Fehling's solution. A 
red precipitate of cuprous oxide will appear. 

(b) With Ammoniacal Silver Nitrate Solution. — Add excess 
of ammonium hydroxide to silver nitrate solution and then 
the liquid containing chloroform. Heat will produce a black 
precipitate of metallic silver. 

These reactions are not characteristic of chloroform, because 
many volatile organic subtances, as formic acid and aldehydes 
which may occur in distillates from animal material, reduce 
Fehling's and ammoniacal silver nitrate solutions. 

Quantitative Estimation of Chloroform in Cadavers 

(Ludwig-Fischer^) 

Mix a weighed portion of material with water and distil 
as long as there is any chloroform. To tell when this point is 
reached, apply the phenylisocyanide test to a few cc. of liquid 

^ An ordinary citrate of magnesium bottle is a convenient apparatus for this 
test. Wrap a towel around the bottle, place it in the water-bath and gradually 
raise the temperature to boiling. Do not remove the bottle until it is cold. Tr. 

' Jahresbericht des chemischen Untersuchungsamtes der Stadt Breslau fdr die 
Zeit vom i April 1894 bis 31 M&rz 1895. 



38 DETECTION OF POISONS 

collected at the end of distillation. Add some calcium car- 
bonate to combine with free hydrochloric acid. Warm the 
distillate to about 60° and draw washed air through it by suc- 
tion. Pass this air through a combustion-tube heated to high 
temperature and then into silver nitrate solution acidified with 
nitric acid. Weigh the precipitated AgCl (N). 

Calculation : 

sAgCl : CHCl, = N : X. 

This method is based upon the fact that chloroform heated 
with steam above 200° is decomposed into carbon monoxide, 
hydrochloric and formic acids: 



i? 



a) CHCl, + H2O = CO + 3HCI. 

) CHCl, + 2H,0 =» H.COOH + 3HCI. 



In a series of blank experiments B. Fischer has shown that the stomach, 
stomach contents and blood, of a person who has not taken chloroform, give 
no volatile chlorine compounds under these conditions. By this method B. 
Fischer found in the cadaver of a laborer, who had died during chloroform 
narcosis, the following quantities of chloroform: 



Weight 


Organ 


Chloroform 


985 grams 


Stomach and contents and parts of 






the intestine 


0.1 gram 


780 grams 


Lungs and blood from the heart 


0-055 g'^am 


445 grams 


Portions of spleen, kidneys and liver 


traces 


480 grams 


Brain 


0.07 gram 



From these results it appears that most of the chloroform was in the brain and 
blood. 

CHLORAL HYDRATE 

CI 

I Chloral hydrate distils very slowly with steam from an add 

I solution. Therefore the complete distillation of a large quantity 

jj Q OH of chloral hydrate requires considerable time. Chloral hydrate 

I appears as such in the distillate. 

OH 

Tests for the Detection of Chloral Hydrate 

Chloral hydrate like chloroform will give the phenyliso- 
cyanide, resorcinol and Lustgarten's naphthol tests. But the 
distillate containing chloral hydrate does not have the charac- 
teristic chloroform odor which is also scarcely perceptible in 
very dilute aqueous chloroform solutions. 



r 



VOLAXILE POISONS 39 



cba 



Jaworowski' suggeats the following tests to diSereatiate 
chloral hydrate from chloroform: 

1 . Test with Nessler's Solution. — Add a few drops of this re- 
agent to an aqueous chloral hydrate solution and shake. It 
will produce a yellowish red precipitate, the color of which will 
change after a while to a dirty yellowish green. This is an 
aldehyde reaction, 

2. Test with Sodium Thiosulphate.^ — Boil a few cc. of chloral 
hydrate solution with 0,2-0.3 gram of solid sodium thiosulphate. 
This will give a turbid liquid of brick-red color. A few drops 
of potassium hydroxide solution will remove the turbidity and 
change the color to brownish red. 

When the quantity of chloral hydrate is not too small, it may 

10 be detected by the following procedure: 

Decomposition of Chloral Hydrate. — Heat a portion of the 
distillate for 30 minutes under a reflux coudenser with calcined 
magnesium oxide (MgO) upon a boiling water bath. Magne- 
sium formate and chloroform are produced by decomposition 
of chloral hydrate. 

iCCl,.CH(OH), + MgO = jCHCii -t- Mg(OOCH)i -|- H.O. 

Proceed as follows to detect these products; 

Chlorofonn.— Distil a few cc. from the solution in the flask 
and test for chloroform by the phenylisocyanide, resorcinol 
and «-naphthol tests. 

Fonnic Acid. — Filter the residue from the distillation, con- 
centrate the filtrate to a few cc, by evaporation and divide into 
two parts for the following reduction tests: 

(a) Reduction of Mercuric to Mercurous Chloride. — Add 
a few drops of mercuric chloride solution and warm. Formic 
add, if present, will produce a white precipitate of mercurous 
chloride (calomel) : 

Mg(OOCH), -I- 4HgCl, - jHg,Cl, + MgCi, + aHCl + aCO,. 

(b) Reduction of Sflver Nitrate.— Wanned with silver nitrate 

' Pharmaceutische Zeilung fur Russlind 33, 37J, und Zeitschritt file aonlylische 
Chemtc, 37, 60 (iSgS). 



40 DETECTION OF POISONS 

solution, formic acid and its salts produce a black precipitate of 
metallic silver: 

Mg(OOCH), + 4AgN0i = 4Ag + Mg(NOi), + 2HNO1 + 2CO,. 

Detection of Chloral Hydrate in Powders or Solutions 

Extract a powder with cold water containing sulphuric add, 
filter, extract the filtrate several times with ether and spon- 
taneously evaporate the ether extracts in a shallow dish or on a 
clock glass. Chloral hydrate imparts to the residue its char- 
acteristic pungent odor. The odor of chloroform is easily 
recognized by warming the residue with sodium hydroxide 
solution: 

CC1,-CH(0H), + KOH = CHCl, + H.COOK + H,0 

The phenylisocyanide, resordnol and naphthol tests, as well 
as that with Nessler's reagent, should be applied to the residue. 

In the case of an aqueous solution of chloral hydrate, first 
acidify with dilute sulphuric add and repeatedly extract with 
ether. jSvaporate the ether extracts and examine the residue as 
already, described. 

Note. — ^ure chloral hydrate forms transparent crystals which are dry, perma- 
nent and colorless. This compound has a pungent odor, its taste being caustic 
and faintly bitter. It dissolves with ease in water, alcohol and ether; and in 5 
parts of chloroform. It melts at 58^. 

Action and Fate of Chlor^ Hydrate in the Hitman Organism 

Applied locally chloral hydrate acts as a strong stimulant. Taken internally 
it frequently stimulates the stomach. When it reaches the blood, it acts like 
chloroform in paralyzing the brain, spinal cord and heart but usually no previous 
stimulation is noticeable. There is marked decrease in blood-pressure due to 
paralysis of the blood-vessels. Death from chloral hydrate poisoning is occa- 
sioned by impaired circulation and respiration, in consequence of which the quan- 
tity of oxygen taken in and of carbon dioxide given off is considerably diminished. 
H. Meyer has shown that the narcotic action of chloral hydrate depends, as does 
that of all compounds of the alcohol and chloroform group, upon the affinity of 
the poison for lipoids, the fatty constituents of the nervous system. It is also 
held by the blood, especially by the red blood-corpuscles. Later it appears 
unchanged, most abundantly in the cells of the brain and spinal cord (Kobert, 
" Intoxikationen ") . 

Only very little chloral hydrate taken internally passes as such into the urine. 
As shown by v. Mering and Musculus,^ the greater part by conjugation with gly- 

^Berichte der Deutschen chemischen Gesellschaft 8, 662 (1875); &i^d v. Mer- 
ing, Ibid., IS, 1019 (1882). 



VOLATItE POISONS 41 

P colonic add forms urochloralic acid (CiHuCIiOt) which is eliminated as such in 
jrinc. This conjugated acid undergoes hydrolysis, when boiled with dilute 
acids, and gives trichlor- ethyl alcohol and free dextro-rotatory glycuronic acid; 
C,Hi,CI,0, + HiO - CCli-CH,OH + H0OC-(CH.0H).-CH0 

■ Urochloralic Trichlor- Glycuronic 

add. ethyl alcohol. add. 

Urochloralic add b therefore trichlor-cthyl glycuronic add. It is crystalline 
mod with heat reduces silver solution as well as alkaline copper and bismuth so- 
lutions. Consequently chlonl urine behaves much like sugar urine but differs 
from the latter in being strongly Itevo- rotatory. The reduction of the aldehyde 
chloral, to its correaponding primary alcohol, trichlor-etbyl alcohol, is especially 

■ noteworthy as regards the behavior of chloral hydrate in the human organism. 
Quantitative Estimation of Chloral Hydrate in Blood and Tissues 
(ArchangelskyO 

Distil the material for ii~io hours with its own weight of 20 per cent, phos- 
phoric add, repeating the process if the distillate is turbid or yellow. To com- 
plete the decomposition of chloral hydrate into chloroform and formic add, add 
50 cc. oE sodium hydroxide solution to the distillate and concentrate on the water 
bath to about jo cc. Neutraliite the solution eiactly and heal for 6 hours on the 
irater bath with an excess of mercuric chloride solution. Finally weigh the 
predpitated mcrcurous chloride. Satisfactory results were obtained by this 
method when known quantities of chloral hydrate were added to blood and 
organs. Using this method Archangelsky has shown that chloral hydrate is not 
uniformly distributed in the blood but is contained especially in the blood-cor- 
pusdes. When narcosis begins there is less chloral hydrate in the brain than in 
the blood. But later the percentage of the poison ia the brain is higher than in 
the blood. Archangelsky his further shown how much chloral hydrate the blood 

I must contain before narcosis can appear. A dog's blood must contain 0.03-0.0; 
percent. When the blood contains 0.1 2 per cent., cespiialion ceases. 
lod 
lo 
HUfi* 



IODOFORM 

Iodoform crystallizes in shining hexagonal leaflets or plates. It may also 
appear as a rather fine crystalline powder, lemon-yellow in color and 
having a penetraring odor somewhat like saffron. The mdting-point 
( iodofonn ia approximately 130°. It is nearly Insoluble in water; 
soluble in 50 parts of cold and in about :o parts o( boiling alcohol; 
and soluble in 6 parts of ether. It is also freely soluble in chloroform. 

Detection of Iodofonn 



T havin 

-C— I of iot 



Iodoform distils quite easily with steam and gives a milky 
distillate having a characteristic odor. Extract this distillate 
with ether and carefully test the residue left by the spontaneous 



■Archiv lur eiperimentelle Palhotogie und Pharroakologie, 46, 347 (looi). 



42 DETECTION OF POISONS 

evaporation of the solvent. If much iodoform is present, 
it will form yellow hexagonal plates. Dissolve the ether 
residue in a little alcohol, and use this solution for the following 
tests: 

I. Lustgarten's^ Test. — Gently warm a few drops of alcoholic 
iodoform solution in a test-tube with a little sodium phenolate 
(CeHs.ONa) solution.^ If iodoform is present, a red substance 
will be deposited on the bottom of the tube. A few drops 
of dilute alcohol will dissolve this precipitate with a carmine- 
red color 

Also make the resorcinol and phenylisocyanide tests (see 
page 36). 

NITROBENZENE 

Nitrobenzene has a strong poisonous action. Administration of very small 

quantities of this compound has produced death in human beings. There are 

-^Q records in the literature of several cases where 30 drops, and even 

r 7 to 8 drops, have caused fatal results. But on the other hand 

. C complete recovery has followed poisoning by much larger doses. 

-^ Y»_ Fatal poisonings have come also from inhaling nitrobenzene vapor. 

ill Within recent years nitrobenzene has been used to some extent as 
CH ^^ abortifadent. Nitrobenzene poisons the blood and changes its 
\y appearance. The blood has a chocolate color and at the same time 
^ the red blood-corpuscles change their shape and go into solution. 

As a result the blood is incapable of uniting with oxygen. The 
blood of persons poisoned by nitro benzene is said to contain less than i per 
cent, of oxygen so that death is caused by asphyxiation. Healthy blood 
contains about 17 per cent, of oxygen by volume. There seems to be no 
methsemoglobin in blood containing nitrobenzene. Such blood examined 
spectroscopically shows the two oxyhsemoglobin bands and also a special 
absorption-band between C and D (Fihlene's nitrobenzene band). It is proba- 
ble that the slight solubility of this poison necessitates a [definite incubation 
period, for 2 to 3 hours usually elapse after nitrobenzene has been taken before 
signs of intoxication appear. A woman, who had taken 10 drops of mirbane oil 
as an abortifadent, gave no indication of intoxication, that is to say, uncon- 
sdousness and cyanosis, for 8 hours after taking the poison. 

Nitrobenzene not only profoundly changes the blood but it irritates and 
paralyzes the central nervous system (see R. Robert, "Intoxikationen")- 

Some nitrobenzene passes into the urine but the organism does not appear to 
convert it into aniline. In nitrobenzene poisoning human urine contains a 

1 Monatshefte fur Chemie, 3, 715 (1882). 

' Prepare sodium phenolate solution by mixing 20 grams of phenol with 40 
grams of sodium hydroxide and 70 grams of water. 



VOLATILE POISONS 43 

brown pigment but only rarely hemoglobin or methemoglobin. Urine contain- 
ing nitrobemsene wiU reduce Fehling's solution. It is also unfermentable and dis- 
tinctly Isvo-rotatory. A conjugated glycuronic acid is possibly concerned in 
these reactions. 

Detection of Nitrobenzene 

In nitrobenzene poisoning the urine and all the organs have 
the odor of this compound. For the chemical tests the material 
should first be distilled with water. Nitrobenzene distils quite 
easily with steam and appears in the distillate as yellowish 
globules. These are heavier than water and have a character- 
istic odor. Vigorously agitate the globules, when separated as 
completely as possible from water, with granulated tin and a 
few cc. of concentrated hydrochloric add, until there is no odor 
of nitrobenzene. Pour the add solution from undissolved tin, 
and add an excess of potassium hydroxide solution to decompose 
the double chloride of aniline and tin. Extract free aniline 
with ether. Withdraw the aqueous liquid from the separating 
funnel, and evaporate the ether extract spontaneously in a 
small glass dish. Aniline, formed by redudng nitrobenzene, 
will remain as globules which usually have a red or brown color. 
Dissolve these globules by agitation with water, and use this 
solution for the hypochlorite and phenylisocyanide tests (see 
pages 45 and 36). 

Mechanism of the Reaction. — ^Nitrobenzene is reduced by nascent hydrogen 
to aniline (a) which combines with the excess of hydrochloric acid forming aniUne 
hydrochloride {fi). From the latter compound potassium hydroxide liberates 
aniline (7): 

(a) CeHi-NOi + 6H = CeH.-NH, + 2H,0, 

05) CHi-NH, + HCl = CeH,-NH,.HClS 

M CiH»NHi.HCl H- KOH = CeH.-NH, + H,0 + KCl. 

^ Organic ammoniimi bases resemble ammonia in combining with adds to form 
salts. Tri\ralent nitrogen of the free base is changed to pentavalent nitrogen in 
the salt: 

* 

III /H V /S 

CeHi = N< H- HCl = C«Hi - NnS 

Aniline Aniline hydrochloride 



44 DETECTION OF POISONS 

ANILINE 

Toxic Action. — ^Aniline is moderately toxic in its action. Doses of 1.5 to a 

grams, administered in the course of a day, have proved fatal to smaU dogs. It 

is not possible to state definitely the average lethal dose for human beings. Very 

-^^ serious results are said to have followed a dose of 3 or 4 grams of 

i aniline. The lethal dose is certainly less than 25 grams, for that 

quantity of aniline was sufficient to kill a healthy man. Even 
Tj(<c^TT I'^^^^O'^ ^^ aniline vapor may cause severe, or fatal intoxications. 

ill Aniline produces methemoglobin and therefore poisons the 

CH blood. The conversion of oxyhemoglobin into methemoglobin 

\^ by aniline may be demonstrated by adding an aqueous aniline 

^ solution to blood in a test-tube. Aniline changes their form and 

partially decomposes red blood-corpuscles. Thereby the quan- 
tity of available oxygen in the blood is so diminished that it amounts to only 5 
to 10 volumes instead of 15 to 20, the normal quantity. The number of red 
blood-corpuscles is diminished in aniline poisoning but not that of the white blood- 
cells. 

R. V. Engelhardt has shown that aniline is partly changed in the human organ- 
ism into aniline black, or into a similar compound insoluble in water. At the 
climax of aniline poisoning blue-black granules may be seen in every drop of blood 
and also in the urine. Aniline is oxidized in the system to para-aminophenol 
(CeH4.0H,NHt(i,4)) . Like all phenols this compoimd forms an ethereal sul- 
phate with sulphuric acid,^ namely, para-aminophenyl-sulphuric add (HO.SOt.- 
O.CeH4.NHt(i,4). This acid is eliminated through the kidneys as an alkali salt 
and then appears in the urine A part of the para-aminophenol is also elim- 
inated as a conjugate of glycuronic acid.' 

The reduction of Fehb'ng's solution by urine containing aniline is due to 
this conjugated acid. In severe cases of poisoning imchanged aniline has also 
been foimd in the urine. Usually urine that contains amline has a very dark 
color. Besides the substances mentioned, a dark pigment has been detected 
in urine in aniline poisoning as well as hsmoglobm, methsmoglobin and an 
abundance of urobilin (R. Kobert, ^'Intoxikationen")* 



Detection of Aniline 

Aniline is a rather feeble base and part of it will pass over 
with steam, when the acid solution is distilled. There will be 
enough in the distillate for detection by the tests described be- 

^ This conjugation takes place with elimination of water: 

H,N.C6H4.0H -f HO.SOj.OH = H,0 + H,N.C«H4.0.S0,.0H (i, 4) 

•Glycuronic add, C6Hio07=Q^C-(CH.OH)4-CC)OH, is closely related to 

glucose. It is an uncrystallizable syrup. If its aqueous solution is boiled, the 
acid is partly converted into the internal anhydride, glycurone (CeHtOi), which 
crystallizes well. 



VOLATILE POISONS 45 

low. In estimating aniline quantitatively in any kind of mate- 
rial the distillation must be as complete as possible. Mix the 
substance with water, make strongly alkaline with sodiiun 
hydroxide or carbonate solution and distil in a current of steam. 
Since 30 parts of water at 15° dissolve i part of aniline, the 
distillate may contain considerable of this amine. When the 
quantity is large, oil-drops will appear. An aqueous aniline 
solution (aniline water) colors pine wood and elder pith 
intensely yellow. The following tests should be used for 
aniline: 

1. Hypochlorite Test — ^Add a few drops of aqueous calcium or 
sodium hypochlorite solution drop by drop to a portion of the 
distillate. A violet-blue or purple- violet color, gradually chang- 
ing to a dirty red, will appear if aniline is present. Addition of a 
little dilute aqueous phenol solution containing some ammonia 
will produce a blue color which is quite stable. This test is 
sensitive i : 66,000.^ 

2. Phenylisocyanide Test. — ^Heat a portion of the distillate 
with a few drops of chloroform and potassium hydroxide solu- 
tion. The repulsive odor of phenylisocyanide will show the 
presence of aniline. 

3. Bromine Water Test — Bromine water added to a solution 
containing aniline will produce a flesh-colored precipitate. 
This test is sensitive i : 66,000. 

4. Chromic Acid Test ^ — Mix a trace of pure aniline with 4 to 
5 drops of concentrated sulphuric acid in a porecelain dish and 
add a drop of aqueous potassium dichromate solution. After 
a few minutes the mixture beginning at the edge will take on a 
pure blue color. Addition of 1-2 drops of water produces at 
once a deep blue color. To apply this test to the distillate, 
first extract with ether, evaporate the ether solution and test an 
oily residue as described. 

* Test this experimentally with very little aniline. For example, dissolve a 
small drop in 30 cc. of water and take only 2-3 cc. of this dilute solution for the 
test. 

' Beissenhirtz reaction, Annalen der Chemie und Pharmazie, 87, 376 (1853). 



46 DETECTION OF POISONS 

CARBON BISULPHIDE 

Carbon disulphide, CSs, is a colorless liquid having a characteristic odor and a 
high index of refraction. It is only slightly soluble in water. There is some 
difference of opinion as regards the solubility of carbon disulphide in water. 

looo cc. of water dissolve 
13-14** 2.03 parts (Page) 

iS-id** 1. 81 parts (Chancel; Parmentier) 

15-16® 2-3 parts (Ckindi) 

15-16** 3.5-4.52 parts (Peligot) 

Carbon disulphide is misdble in all proportions with absolute alcohol, ether, 
ethereal and fatty oils. 

Toxic Action. — Carbon disulphide administered internally has a very poisonous 
action upon the blood causing especially decomposition of red blood-corpusdes. 
Even inhalation of carbon disulphide vapor frequently occasions severe poisoning. 
Carbon disulphide was formerly considered a typical producer of methsmoglobin 
but recent investigations have not confirmed this opinion. It has a very injurious 
action upon the red blood-corpuscles and causes hemolysis. R. Robert (Intox- 
ikationen) states that its power of dissolving lipoids is responsible for its injuri- 
ous action upon the blood and the central nervous system. E. Harmsen^ has 
recently come to practically the same conclusion. He considers carbon disul- 
phide a powerful blood poison because it decreases the nimiber of red blood-cor- 
puscles and the quantity of hemoglobin and brings about a leucocytosis.' 

Detection of Carbon Disulphide 

Carbon disulphide distils very slowly with steam. Con- 
sequently the second or third fraction of the distillate should 
be used in testing for this substance. If 40 cc. are distilled 
from 100 cc. of water containing 2 drops of carbon disulphide, 
the following 10 cc. will give a distinct test. If the quantity 
of carbon disulphide is small, it will remain in solution. Such 
a solution does not have a strong odor. Carbon disulphide may 
be recognized by the following tests: 

I. Lead Acetate Test. — ^Add a few drops of lead acetate 
solution to the liquid containing carbon disulphide. It will 
cause neither a precipitate (distinction between CS2 and HiS) 
nor a color. Add excess of potassium hydroxide solution and 
boil. A black precipitate (PbS) will appear. This is a very 
delicate test. 

^ Vierteljahrsschrift fUr gerichtliche Medizin, 30, 422 (1905). 

* Leucocy tosis means a temporary increase in the number of white blood- 
corpuscles (leucocytes) as compared with the number of red blood-corpuades. 
Normally there are about 350 red to i white blood-corpuscle, whereas in leucocy- 
tosis the proportion is 20 : i. 



VOLATILE POISONS 47 

2. Sulphocyanate Test. — Heat an aqueous solution of carbon 
disulphide for a few minutes with concentrated ammonium 
hydroxide solution and alcohol. Ammonium sulphocyanate 
(H4NSCN) is formed together with ammonium sulphide. 
Concentrate this solution upon the water-bath to about i cc. 
and acidify with dilute hydrochloric acid. Add a drop of 
ferric chloride solution and a deep red color will appear. This 
test will show even traces of carbon disulphide, for example 
0.05 gram in i cc. of water. 

Mechanism of the Reaction: 

(a) 4NH, + CS, = (H4N)SCN + (H4N),S, 

O) FcCU + 3(H4N)SCN - (Fe(SCN), + 3(H4N)C1. 

3* Xanthogenate Test — Shake a few cc. of distillate for 
several minutes with 3 or 4 times its volume of saturated solu- 
tion of potassium hydroxide in absolute alcohol. Faintly 
acidify the solution with acetic add and add i or 2 drops of 
copper sulphate solution. If carbon disulphide is present, a 
brownish black precipitate of cupric xanthogenate will appear. 
This will soon change to a yellow, flocculent precipitate of 
cuprous xanthogenate, CS(SCu) (OC2H5) . Vitali's procedure is 
somewhat different and consists in adding solid ammonium 
molybdate to the alkaline reaction-product and then in acidify- 
ing with dilute sulphuric add. The appearance of a red color 
indicates carbon disulphide. 

Mechanism of the Reaction. — Alcoholic potassium hydroxide acts like 
potassium alcoholate (CtHt-OE) and converts carbon disulphide. in to potassium 
xanthogenate 

/SK 
CS, + CtHi-OK - C==S 

\OC,Hi 

This compound treated with cupric salts gives first a brownish black precipitate 
of cupric xanthogenate: 

/SK /S— 

2C==S H- CuSO* = (S = C< ),Cu H- K,S04 

\0C,H| X)C,H, 

The cupric salt then forms cuprous xanthogenate and ethyl xanthogen disulphide : 

<0CjH6 /OCjHi yOCHi 

S = C< S = C< 

Sv ^S ^S - Cu 

2 >cu - I + r 

/$/ yS yS - Cu 

s = c< s = c< s = c<; 



OC,Hi ^OC,H» ^OC,Hi 

Cupric Ethyl xanthogen Cuprous 

xanthogenate disulphide xanthogenate 





CSa in mgrs. 




per liter of air 


I. 


0.5-0.8 


2. 


1.3 


3. 


3.4 


4. 


6.0 


5. 


10. c 



48 DETECTION OF POISONS 

Quantitative Estimatioii of Carbon Disulphide In Air 

Inhalation of air containing carbon disulphide has frequently given rise to 
chronic poisoning. Persons thus affected have usually been laborers in rubber fac- 
tories. Consequently experiments have been made to determine the maximum 
quantity of carbon disulphide air may contain without injury to health. The 
results of these investigations may be summarized as follows: 

Result 

No injurious effect. 

Slight uneasiness after several 
hours. 

Uneasiness in 30 minutes. 

Uneasiness in 20 minutes. 

Paralysis attended by after-effects last- 
ing several days. 

The exact danger limit for persons obliged to live for weeks at a time in an atmo- 
sphere containing carbon disulphide should be placed below 3 mg. per liter 
of air. Air in factories, where operatives work in presence of carbon disulphide 
vapor, should never exceed this limit. In rubber factories the air is said fre- 
quently to contain 2.5 to 3 mg. per liter. Since experiments have shown 
that 93 to 96 per cent, of the carbon disulphide breathed was exhaled imchanged, 
an exceedingly small quantity is capable of producing toxic symptoms. 

Procedure. — Place a saturated alcoholic solution of potassium hydroxide in a 
P^ligot absorption- tube and draw through this solution 10 to 20 liters of air con- 
taining carbon disulphide vapor. A quantitative formation of potassium zan- 
thogenate (see above) will take place. 

Dilute the contents of the receiver at the end of the experiment with 96 per 
cent, alcohol and bring the volume to 50 cc. Measure an aliquot portion of this 
solution and dilute with water. Faintly acidify the solution with acetic add 
and remove excess of add with add sodium carbonate. Add freshly prepared 
starch solution and o.i n-iodine solution until there is a permanent blue color. 

Iodine converts potassium xanthogenate according to equation (I) into ethyl 
zanthogen-disulphide : 

KS.CS.OCHi S.CS.OCHi 

I. It H- - 2^1 + T 

KS.CS.OCH, S.CS.OCHi 

£. Rupp and L. Krauss^ think the action of iodine upon potassium xanthogen- 
ate is expressed by equation (II) : 

II. 2KS.CS.0C,H. -f H,0 H- 2I = KS.CS.sk + 2C2H».OH + 2HI + S. 

Both equations require the same quantity of iodine, namely, 2 atoms for 2 
molecules of xanthogenate. A difference therefore in the mechanism of the 
reaction has no influence on the combining relations of the iodine and the method 
is applicable to the quantitative determination of xanthogenate. 

1000 cc. of o.i n-iodine solution, containing o.i gram-atom of iodine, corre- 
spond to 0.1 gram-molecule of CSa =7.6 grams. 

^ Berichte der Deutschen chemischen Gesellschaft 35, 4257 (1902). 



VOLATILE POISONS 



ETHYL ALCOHOL 



f Fate in the Human Orgknism. — Alcohol brought in contact with m&ay ilSer- 

felt parts of ihc organism is very rapidly absorbed, but especially easily from an 

"empty atomadi. Although there is practically no absorption of non-voUtlle 

„ aqveoiu tiquids from the stomach, alcohol is freely absorbed. 

I After absorption it passes into the blood and is then distributed 

H — C — H toallorgaDB (see chloral hydrate). Experiments upon dogs, colls 

J.^j-iTi '■'"' aduit horses (see Robert, "Intonkatioaea") have shown 
H— C— OH j),n( ^[jjjjj ^j j^g climai of narcosis contains 0.7* per cent, of 

H alcohoL There is stupor even when c.ii per cent, is present. 

There is diScrence of opinion among toxicologista regarding 
alcoholic intoxication, as to whether the poison is distributed unifornJy through- 
out the body, or accumiUated in the brain in larger quantity than in other 
organs. The following percentages of alcohol, found in the organs of a man, 
tcho had died at the climax of severe acute alcohol poisoning, lead support to 
the latter view; liver o.ti, brain 0.47 and blood 0.33 per cent. It appears from 
these results that the brain takes up an especially large quantity of alcohol. 

Uncertainty concerning the subsequent fate of alcohol in the organism has 
finally been removed. Experiments have shown that alcohol is never eliminated 
unchanged through the skin. .\t most only 1-1.5 P*'' cent, passes off through 
the kidneys and only i ,6~i per cent, througbt the lungs. Strassmaim' found the 
quantity eliminated by the lungs somewhat higher (5-6 per cent.) and by the 
kidneys i-J-s per cent. The remainder is completely oxidized In the human 
organism to carbon dioxide and water. 

B. Fischer found the following quantities of alcohol in organs removed from a 
mAn who had probably died from drinking too much brandy : 



Weight 


Organ 


Alcohol 


3710 grams 


Stomach and intestines 


30.6 grams 


3070 grams 


Heart, lungs and blood 


10.85 grams 


1830 grama 


Kidneys and Liver 


7,8 grams 


136s grams 


Brain ' 


4.3 grams 



r Detection of Ethyl Alcohol 

Ethyl alcohol distils easily with steam and consequently 
most of it will be in the first fraction. If present in sufficient 
quantity, it can be recognized in the distillate by its odor. The 
following tests should be made: 

I. Lieben's Iodoform Test.* — Gently warm the hquid (40- 

50°), add a few cc. of aqueous iodo-potassium iodide solution, or 

_a small crystal of iodine, and enough potassitmi hydroxide 

olution to give the liquid a distinct yellow to brownish color. 

' Pfiiiger'E Archiv, 49, 315 (1S91). 

' Annalen der Chemie und Pharmazie, Supplement Band, 7, iiS. 



50 DETECTION OF POISONS 

If alcohol is present, a yellowish white to lemon-yellow precipi- 
tate of iodoform will appear immediately, or as the solution 
cools. If the quantity of alcohol is very small, a precipitate will 

form on long standing. When iodo- 
form is deposited slowly, the crystals 
are very perfect hexagonal plates and 
stars (see Fig. 9). 




Note. — ^This iodoform test is very delicate 
but not characteristic of ethyl alcohol, since 
other primary alcohols, except methyl alcohol, 
and many secondary alcohob, as well as their 
oxidation products, aldehydes and ketones, 
give iodoform imder the same conditions. 

Fig. 9.— Iodoform Crystals. ^^^^ ®^^» aceto-acetic ether, lactic add. etc, 

also give iodoform. 
The correct explanation of the iodoform reaction is probably the following: 
Iodine and potassium hydroxide form potassium hypo-iodite (KOI) by reaction 
(a). This compoimd brings about the oxidation of alcohol to acetic aldehyde 
(fi) and at the same time substitutes iodine for hydrogen in the latter (y ) . Finally 
tri-iodo-acetic aldehyde is decomposed by the excess of potassium hydroxide into 
iodoform and potassium formate (5) : 

(a) 2KOH H- I, = KIH- H,0 + KOI, 

03) CH,.CH,.OH + KOI = CH,.CHO + H,0 + KI, 
(7) CH,.CHO + 3KOI » 3KOH + CI1.CHO, 
(6) CI,.CHO H- KOH = CHIi + H.COOK. 

2. Berthelot's Test — Shake the liquid containing alcohol 
with a few drops of benzoyl chloride and about 5 cc. of sodium 
hydroxide solution (10 per cent.), until the irritating odor of 
benzoyl chloride has gone. The aromatic odor of ethyl ben- 
zoate will appear. 

C«H6.C0C1 H- C,H,.OH + KOH = C6H,.C0.0C,H, + KCl + H,0 

Ten cc. of 0.5 per cent, alcohol will give a distinct odor of this 
ester. 

3. Chromic Acid Test. — ^Warm the liquid containing alcohol 
with dilute sulphuric, or hydrochloric acid, and add i or 2 drops 
of very dilute potassium dichromate solution. The color of the 
liquid will change from red to green, and at the same time the 
odor of acetaldehyde will be recognized. This test is not char- 
acteristic of alcohol, because many other volatile organic 
compounds behave similarly. 



VOLATILE POISONS 51 

Mechanism of the Reaction 

(a) KiCrjOy + H,S04 - K1SO4 + H,Cr,07(H,OH-2CrOi), 

(/3) 3 C»H..OH + 2 CrO, + 3 H,S04 = 3 CH,.CHO + Cr,(S04)iH- 6H,0. 

Acetaldehyde 

4* Ethyl Acetate Test — Mix the liquid containing alcohol 
with the same volume of concentrated sulphuric add. Add a 
very small quantity of anhydrous sodium acetate and heat. 
Ethyl acetate will be recognized by its odor. 

(a) C1H1.OH + H1SO4 - CHjO.SCOHi + H,0, 

O) CH,.CO.ONa + C,HiO.SO,.OH = CHi.CO.OCiHi + NaHSO*. 

5. Vitali's Test. — Thoroughly mix a few cc. of distillate in a 
glass dish with a small piece of solid potassium hydroxide and 
2 or 3 drops of carbon disulphide. Let this mixture stand for a 
short time without warming. When most of the carbon disul- 
phide has evaporated, add a drop of ammonium molybdate 
solution and then an excess of dilute sulphuric acid. If the dis- 
tiUate contains alcohol, a red color will appear. Potassium 
xanthogenate (CS(OC2H6)(SK)) is first formed. This com- 
pound gives a red color with ammonium molybdate. Acetone 
and acetaldehyde produce a similar color. This test is given 
distinctly by s per cent, aqueous alcohol solution. 

ACETONE 

Human urine almost always contains a very small quantity of acetone as a 

physiological constituent. Under pathological conditions, especially in diabetes 

U mellitus (diabetic acetonuria), urine contains much more. It is 

I also present in urine in protracted high fever, digestive disturb- 

"^ — C — ^H ances, severe forms of carcinoma (carcinomatous acetonuria), etc. 

, ^ Finally, acetone has been found in urine in considerable quantity 

A' in various intoxications (toxic acetonuria), for example, in poison- 

— ^H ing by phosphorus, carbon monoxide, atropine, curare, antipyrine, 
pyrodine, sulphuric acid, extract of male fern; in chronic lead 
poisoning; and in chronic morphinism after discontinuance of the 
drug (see R. Kobert, "Intoxikationen"). 

Acetone is not poisonous nor in the least corrosive. Man and animals can 
tolerate considerable quantities of acetone taken internally. It seems to produce 
no effect, though it may possibly possess very feeble narcotic properties. Axch- 
angelsky found dogs to show signs of narcosis when the blood contained 0.5 per 

XOCaHi 
OH 






52 DETECTION OF POISONS 

cent, of acetone. Even smaller doses produce narcosis in rabbits and have an 
injurious action upon the blood and kidneys. 

Distillates from human urine, as well as from blood and various organs, as 
liver, spleen, kidneys, brain, etc., often contain acetone, or more correctly per- 
haps, substances like acetone. This is especially the case when cadaveric mate- 
rial has begim to putrefy. 

Acetone is a clear, colorless liquid boiling at sd**. It has a peculiar, fruity odor 
and is neutral in reaction. It is misdble in all proportions with water, alcohol 
and ether. It distils easily with steam. 

Detection of Acetone 

1. Lieben's lodofonn Test — ^Add a few cc. of aqueous lodo- 
potassium iodide solution, or a small crystal of iodine, to an 
aqueous solution of acetone and then potassium hydroxide 
solution drop by drop until the color is yellow. Iodoform 
immediately separates, even in the cold, as a yellowish white 
precipitate which is usually amorphous. Acetone differs from 
alcohol in giving iodoform, when ammonium hydroxide solution 
is substituted for potassium or sodium hydroxide solution 
(Gunning's acetone test). 

Acetaldehyde resembles acetone in giving iodoform in the cold 
and under conditions the same as those stated above. 

Note. — Potassium hypo-iodite (a) probably converts acetone into tri-iodo- 
acetone (CH|.CO.CI«) (/3) and this compound is then decomposed by potassium 
hydroxide into iodoform and potassium acetate (7) : 

(a) 6K0H + 3I1 = 3KI + 3KOI + 3H,0, 

03) CH,.CO.CH, + 3KOI = CH,.CO.CI, + 3KOH. 

(7) CH,.CO.CI, + KOH = cm, + CH,.CO.OK. 

2. Legal's Test. — ^Add a few drops of freshly prepared sodium 
nitroprusside solution to a Uquid containing acetone, and then 
potassium hydroxide solution. A red or reddish yellow color 
will appear. This color soon changes to yellow. Add an 
excess of acetic acid to the solution. The solution will now 
have a carmine to purphsh red color, according to the quantity 
of acetone present. Heat will change this color to violet. 

Alcohol does not give Legal's test, though acetaldehyde does. The red color 
caused by aldehyde fades upon addition of acetic add, and changes to green with 
heat. Le Nobel states that ammonium hydroxide, or ammonium carbonate solu- 
tion, may be substituted for potassium hydroxide solution in Legal's test, but 
under these conditions the red color is very slow to appear. Le Nobel's modifica- 
tion, however, eliminates the possibility of confusing acetone with acetaldehyde. 



VOLATILE POISONS 53 

3. Penzoldt's Test. — Prepare a hot, saturated, aqueous 
ortho-nitTO-benzaldehyde (CBHi.N0i.CH0(i,2)) solution and 
allow it to cool. Add this solution to the liquid containing 
acetone, and also some sodium hydroxide solution. At first 
the color of the mixture is yellow. It then becomes green, and a 
blue precipitate of indigotin is formed in lo to 15 minutes. 
When indigotin is present in traces only, shake the solution with 
chloroform. This solvent will dissolve the coloring matter and 
become blue. 

4. Reynold's Test.— Acetone will dissolve freshly precipi- 
tated mercuric oxide, and this test is based upon this property. 
Add mercuric chloride solution to the distillate, and an alcoholic 
potassium hydroxide solution. Shake thoroughly and filter. 
Add ammonium sulphide solution to the clear filtrate as an upper 

J layer. If acetone is present, there will be a black zone (HgS) 
fwhere the two solutions meet. 



■ DetectJOD of Acetone iaDrine. — Acidify 300 to 50a cc. of urine with a few drops 
of sulphuric add and distil. Collect 90 to 30 cc. of disIiUate, This will contain 
the entire quantity of acetone in the urine. Acetone thus oLtained may 

^ possibly be derived from acelo-acetic acid which is often present in human 
urine, especially in a severe case of diabetes mellitus. Dbtillation decomposes 
tliis add into acetone and carbon dioxide. 
CH,.CO.CH,.C0.OH = CHi.CO.CH, + CO,. 

Detection of Alcohol and Acetone in Mixtures. — Alcohol may be detected in 

presence of acelone by Berthelol's test. On (he other hand, acetone may be 
distinguished from alcohol by Lcgal'sot Penzoldt's teat. 



^Btnac 
^fthis 



BITTEH ALMOND WATER AND BENZALDEHYDE 

Bitter almond water (Aqua Amygdalfe Amarse of the Phar- 

lacopceia) contains hydrocyanic acid. Only a small portion of 

acid, however, is free so that it can be precipitated by silver 

nitrate solution. The greater part is combined as the cyano- 

hydrin of benzaldehyde, Cdfi.c^OH, which does not react with 

silver nitrate, But potassium hydroxide solution will decompose 
this compound. 

CiHiCH (OH) CN + KOH - KCN + H,0 + C1H..CHO . 



54 DETECTION OF POISONS 

Pure benzaldehyde, also called hydrocyanic acid-free oil of 
bitter almonds, is not poisonous. It is oxidized to benzoic acid 
in the body and eliminated in the urine partly as that acid and 
partly as hippuric acid after conjugation with glycocoU (amino- 
acetic acid) : 

(a) C«H».CHO +0 - CeH»-COOH, 

03) C«H»-COOH -f H,N-CH,-COOH = C.H»-CO.NH-CH,-COOH. 

Benzoic acid GlycocoU fttrnished Hippuric acid 

by the organism 

Ordinary commercial oil of bitter almonds contains hydro- 
cyanic acid and is poisonous in proportion to the quantity of this 
acid present. 

Test for hydrocyanic acid by shaking about 2 cc. of oil of 
bitter almonds with 20 cc. of potassium hydroxide solution and 
making the Prussian blue test. When oil of bitter almonds is 
mixed with other material, distil with steam from a solution 
acidified with tartaric, or dilute sulphuric acid, and test the first 
part of the distillate for hydrocyanic acid. K benzaldehyde is 
present, the distillate at the same time will be milky and have 
the characteristic odor of that compound. Distil imtil the drops 
of water are perfectly clear. Benzaldehyde may be detected 
with certainty, and at the same time distinguished from nitro- 
benzene which has a somewhat similar odor, by adding a few 
drops of potassium hydroxide solution to the milky distillate, 
to combine with any hydrocyanic acid, and extracting with 
ether. The ether upon evaporation will deposit benzaldehyde 
as globules, which can be positively identified by conversion into 
benzoic acid. Heat the globules for a few minutes in a small 
flask, attached to a reflux condenser, with about 10 cc. of potas- 
sium dichromate solution and a little dilute sulphuric add. 
Cool, extract with ether and evaporate the ether solution in a 
glass dish. When the material contains benzaldehyde, tTiig 
residue will consist of benzoic acid. This substance may be 
further identified by its melting point (120-121°), its property 
of subliming and the test with ferric chloride solution.^ 

^ Dissolve the residue in a small quantity of water, and neutralize benzoic add 
by heating the solution to boiling with excess of calcium carbonate. Filter and 
add a few drops of ferric chloride solution. If benzoic add b present, a fleali- 
colored precipitate of basic ferric benzoate will appear. Tr. 




VOLATILE POISONS 



SYNOPSIS OF GROUP I 



I 



ScheTer*! Test for Phosphorus Precedes Distillatton 

The material to be examined must first be rendered unifonn 
by grinding or chopping. Add sufficient water to thin the mass, 
acidify with tartaric acid and distil. If the preliminary test 
for phosphorus (Scherer's) is positive, distil in the Mitscherlich 
apparatus; otherwise distil as usual with a Liebig condenser. 
It is advisable to collect the distillate in two or three fracrions. 
Test the first 5 to 10 cc. of distillate for hydrocyanic add, 
chloroform, ethyl alcohol, acetone and possibly also for nitro- 
benzene and iodoform. Use the remainder of the distillate 
in testing for carbolic add, chloral hydrate and carbon 
disulphide. 

I^OBphorus. — Phosphorescence in Mitscherlich apparatus 
during distillation in a dark room. Evaporate distillate with 
strong chlorine water, or a little fuming nitric add, and test the 
residue for phosphoric acid. As an alternative procedure, 
examine the original material, or at least the Mitscherlich dis- 
tillate, for phosphorus by the BIondlot-Dusart method. 

Hydrocyanic Acid.^ — Odor. Schdnbein's preliminary test. 
Prussian blue test. Sulphocyanate test. Nitroprusside test. 
Silver nitrate test. Alkaline phenolphthalin test. 

Carbolic Add. — Odor. Red color with Millon's reagent. 
Yellowish white precipitate with bromine water. Violet color 
with ferric chloride solution. 

Chloroform. — Separation of colorless globules, when the 
quantity is large. Odor. Phenylisocyanide test, when heated 
with aniline and potassium hydroxide solution. Reduces silver 
nitrate and Fehling's solutions with heat. Red color with 
resorcinol and potassium hydroxide solution. Blue color with 
naphthol and potassium hydroxide solution. 

Chloral Hydrate. — Gives chloroform reactions. Brick-red 
predpitate with Nessler's solution which in time becomes 
yellowish green. Gives chloroform and magnesium formate, 
when heated with magnesium oxide and water. Test for 
ftgtnate with silver nitrate or mercuric chloride solution. 



56 DETECTION OF POISONS 

lodofonn. — Odor. Distillate milky and yellowisli white. 
Ether extract of distillate leaves crystals upon evaporation, 
gives chloroform reactions. 

Nitrobenzene. — Yellowish globules with characteristic odor. 
Reduced to aniline, when shaken with tin and hydrochloric 
acid. Test for aniline. 

Aniline. — ^Violet color with calcium hypochlorite solution. 
Phenylisocyanide test, when heated with chloroform and 
potassium hydroxide solution. Flesh-colored precipitate with 
bromine water. Dark red color on warming with Millon's 
reagent. 

Carbon Bisulphide. — Black precipitate, or only black colora- 
tion (PbS), when heated with lead acetate and potassium 
hydroxide solutions. Formation of ammonium sulphocyanate 
by evaporation with concentrated ammoniimi hydroxide solu- 
tion and detection with ferric chloride solution. Formation 
of potassium xanthogenate, when shaken with alcoholic solu- 
tion of potassium hydroxide and detection with copper sulphate 
solution. 

Ethyl Alcohol. — Iodoform test. Odor of ethyl benzoate, 
when shaken with benzoyl chloride and sodium hydroxide solu- 
tion. Green color and aldehyde odor, when heated with potas- 
sium dichromate and hydrochloric acid. Vitali's test. 

Acetone. — Gives iodoform, even in the cold, with iodine 
and potassium hydroxide or ammonium hydroxide solution. 
Legal's'test. Indigotin test. Reynold's test. 



CHAPTER n 
NON-VOLATILE POISONS^ 

Alkaloids, Ghicoddes and Synthetic Compounds Non-volatile from Add 

Solution with Steam 

Put a portion of finely chopped material into a large flask, 
and thoroughly mix with two or three times as much absolute 
alcohoL* Add enough tartaric acid solution to give the mixture 
a distinct acid reaction after shaking. Laboratory experiments 
usually require 20 to 30 drops of 10 per cent, tartaric acid solu- 
tion. Avoid a large excess of tartaric acid, since it may act 
as an objectionable impurity in the ether extract, owing to its 
solubility in that solvent. Connect the flask with a glass tube 
(80 to 100 cm. long) serving as a reflux cooler. Frequently 
shake and heat 10 to 15 minutes upon the water-bath. In the 
extraction of a large quantity of material from a cadaver, con- 
nect the flask with an upright Liebig condenser used as a reflux 
cooler (Fig. 10). Cool the flask contents and filter to remove 
fat and other insoluble matter. Wash the residue with alcohol. 
Evaporate the filtrate, which must have an acid reaction, to a 
thin syrup in a glass dish upon the water-bath. Thoroughly 
mix this residue with 100 cc. of cold water. Usually this 
causes an abundant separation of fat and resinous matter, 
especially when parts of a cadaver are examined. Filter and 
evaporate the filtrate to dryness, or to a syrup, upon the water- 

^ The isolation of these toxic substances from cadaveric material, food, etc., 
is necessary before tests establishing their presence can be made. Mixtures used 
for laboratory practice, consisting of dog biscuit, meat, comminuted organs (liver, 
kidneys, spleen), sausage meat, etc., with any of the poisons of this group, should 
be examined according to the method outlined above. 

* Commercial alcohol usually contains basic compounds, the presence of which 
is objectionable. They should be removed by adding tartaric add to the alcohol 
and distilling. Alcohol should not be used in toxicological analysis, imless an 
actual test has shown it to be free from such impurities. Tr. 

57 



58 



DETECTION OF POISONS 



3 



df 



bath. Thoroughly mix this residue with absolute alcohol. As 
a result of this treatment, a whitish substance, which is more or 
less viscous or slimy, usually remains undissolved. Hiis resi- 
due, which consists chiefly of protein substances (albumin, 
albumoses and {>ep tones), dextrin-like compoimds and in part 

also of inorganic salts, frequently be- 
comes granular upon standing. Tartrates 
of the alkaloids and other organic poisons 
are dissolved. The larger the quantity 
of absolute alcohol used, the more com- 
plete the precipitation of those substances 
which interfere more or less with the detec- 
tion of organic poisons. Again evaporate 
the filtered alcoholic solution upon the 
water-bath, and dissolve the residue in 
about 50 cc. of water. If the solution is 
not {>erf ectly dear, filter through a moist- 
ened pa|>er. 

The result of this procedure is a solution 
containing alkaloidal tartrates and other 
organic substances belonging to this group. 
This solution should have an add reac- 
tion and be practically free from protein 
substances, fat, resinoxis bodies and color- 
ing matter. If the solution fulfils these 
requirements, it is ready to be examined 
for organic poisons according to the " Stas- 
Otto^* method. The utmost care must be 
taken in preparing this solution, because 
definite condusions cannot be drawn from 
the uncertain tests given by impure material. 

When the material is a innvder mixed with cane- or milk- 
sugar, it is usually possible* after the aqueous solution has been 
acidineil with tartaric acid» to extract directly with ether and 
continue accoriling to the Stas-Otto method. 

Frequently in suspected jnusoning an examination of beer, 
wine, black cotTee» infusion of tea, t\HHl, etc., is necessary. In 




Fig. xo. — liebig Con- 
denser as Reflux CcK^ler. 



NON-VOLATILE POISONS 59 

''Buch cases the process outlined above may often be considerably 
shortened. Acidify the material with aqueous tartaric acid 
solution, if necessary-, and evaporate in a glass dish upon the 
water-bath. Treat the residue thoroughly with absolute alcohol 
and filter. Evaporate the filtrate upon the water-bath and 
dissolve the residue in tepid water. Filter this solution, if 

BSiecessary, and then examine according to the Stas-Otto'process. 




— Separating Funnels and Glass CryslallJMOg Dishes. 

STAS-OTTO PROCESS 
lation of Ether Extract of Tartaric Acid Solution 
Thoroughly extract the acid aqueous solution (see process of 
preparation described above) two or three times with ether, 
■ using each time about the same quantity of solvent. Employ 



60 DETECTION OF POISONS 

a separating funnel for this purpose (Fig. ii). Pour the com- 
bined ether extracts into a dry flask loosely stoppered. If the 
solution stands for i or 2 hours at rest, a few drops of water 
usually settle out. Decant the ether solution and pour through 
a dry filter. Slowly evaporate this solution in a small glass 
dish upon a water-bath previously heated slightly above 35^ 
Do not have gas burning during this operation! Examine the 
residue as described below. An excellent method of evaporat- 
ing ether consists in setting a small glass dish (8 to 10 cm. in 
diameter) upon a hot water-bath and dropping the filtered ether 
extract into it as fast as the solvent evaporates. Thus a large 
quantity of extract may be evaporated in a small dish. The 
advantage of this method is the ease with which the residue can 
be removed for the various tests. The residue is usually quite 
small and it is not advisable to have it distributed over too 
large a surface. 

Examine the residue from the ether extract for the following 
substances: 

Picrotoxin Caffeine Antipyrine 

Colchidn Acetanilide Salicylic Add 

Picric Add Phenacetine Veronal 

Evaporation of the ether extract, even in the absence of 
members of the group, usually leaves a more or less viscous 
residue, containing tartaric and lactic acids as well as fatty, 
resinous and colored substances. This is espedally so in analy- 
ses of cadaveric material. Moreover ether extracts from aque- 
ous solutions certain metallic salts, for example, mercuric 
cyanide^ and chloride. 

^ Ether will extract meroiric cyanide to some extent from a tartaric add solu- 
tion which is not too dilute. For instance, it will remove appredable quantities 
from 100 cc. of o.i per cent, merouic cyanide solution, but the extraction will not 
be complete. The solution after five extractions will still give a distinct test for 
merciuy. Ether will not remove even a trace of mercuric cyanide from 0.01 per 
cent, solution. To test for cyanide, add ammonium sulphide solution to the 
ether residue. This will predpitate mercuric sulphide and the filtrate will con- 
tain ammonium sulphocyanate (see hydrocyanic add, page 22). 



NON-VOLATItE POISONS 



61 



First, note the general appearance and taste of the residue. 
Then examine it with a microscope. Very definite conclusions 
as to the presence or absence of certain substances can frequently 
be drawn. A very bitter residue should be examined carefully 
for picrotoxin and colchicin. If there is a pronounced yellow 
color, the examination should include picric acid also. Veronal 
is colorless and has a very bitter taste. A tasteless, or only 
faintly bitter, residue probably does not contain these sub- 
stances and should be examined for acetanUide, antipyrine, 
caffeine, phenacetine and salicylic acid. 

The residue from evaporation of the ether extract may con- 
tain the following substances: 

Picrotoxin. — Usually a thick syrup which gradually solidifies 
and becomes crystalline. Tastes intensely bitter. 

Colchicin.— Yellowish, amorphous residue which does not 
become crystalline. Tastes intensely bitter. Dissolves in 
water with a yellowish color, which increases in intensity on 
addition of a few drops of dilute hydrochloric acid. 

Picric Acid. — Usually appears as a syrup which gradually 
solidifies and becomes crystalline. Tastes very bitter. Resi- 
due intensely yellow, giving yellow aqueous solutions not in- 
tensified by hydrochloric or sulphuric acid. 

Acetamlide. — Leaflets or flattened needles. Has a faint, 
burning taste but is not bitter, 

Phenacetine.' — Inodorous and tasteless leaflets and small 
needles, 

Antipyrine. — Residue a syrup which is rarely crystalline. 
Tastes mildly bitter. Very easily soluble in water. 

Caffeine. — Residue composed of shining needles frequently 
in radiating clusters. Tastes mildly bitter. 

Salicylic Acid. — Crystallizes frequently in long needles. 
Tastes harsh and at the same time sweet and acid. 

Veronal. — Crystalline needles having an agreeable, bitter 
^ taste. 

PICROTOXIN 
igHjiOii, the poisonous principle of Cocculus indicos, ihe fruit of 

Dispennum Cocculua, crystallizes from hot water in long colorless needles 



62 DETECTION OF POISOKS 

melting at 1 99-200**. It dissolves with difficulty in cold water but more readily in 
hot water or alcohol. It is slightly soluble in ether but freely soluble in chloro- 
form, amyl alcohol and glacial acetic acid. Its alcoholic solution b neutral and 
IflBvo-rotatory. Picrotozin has a very bitter taste. It is not as readily soluble in 
adds as in pure water, but is soluble in caustic alkalies and aqueous ammonia, 
forming unstable, salt-like compounds which do not oystallize. Picrotozin 
behaves toward strong bases as if it were a weak add. Heated to boiling with 
twenty times its volume of benzene, it is decomposed into picrotozinin and picro- 
tin. The former passes into solution but picrotin is almost completely insoluble: 

CioHt40it ■■ CiiHiiGc -h CiiHiiGt 

Picrotozin Picrotoxinin Picrotin 

Chloroform brings about this deavage even more easily. On the other hand, 
if picrotoxinin and picrotin in molecular proportions are dissolved in hot water, 
picrotozin crystallizes out as the solution cools. Treated with bromine direct or 
dissolved in water or ether, picrotozin is first split into picrotozinin and picrotin. 
The former is immediately converted into monobromo-picrotozinin, Ci»Hi»BrO«, 
but picrotin remains almost unchanged. Monobromo-picrotozinin is soluble 
with difficulty in water but b reduced by zinc dust and acetic add to picrotozinin. 
Picrotin is almost non-toxic, whereas picrotozinin has a very poisonous action. 
Picrotozin is a powerful convulsive poison, standing in its action between dcu- 
tozin and strychnine. 

R. Meyer and P. Bruger^ regard picrotozin as a complez of the two compounds, 
picrotin and picrotoxinin, crystallizing together in definite but not molecular 
proportion, and not as a molecularly constituted chemical compound. 

Detection of Picrotoxin 

1. Fehling's Test — ^Dissolve picrotoxin in a small test-tube, 
using 10-20 drops of very dilute sodium hydroxide solution. 
Add a few drops of Fehling's solution^ and warm but do not 
shake. A red or yellowish red precipitate forms and settles to 
the bottom. If the ether residue, not too little of which should 
be taken, fails to give a clear solution in very dilute sodium 
hydroxide solution, filter through moistened paper and examine 
the filtrate with Fehling's solution. 

2. Ammoniacal Silver Test. — Warm picrotoxin with aqueous 
silver nitrate solution containing a slight excess of ammonium 
hydroxide solution. The reducing action of picrotoxin will 
produce a black precipitate of metallic silver, or a dark brown 
color when only traces are present. 

^ Berichte der Deutsehen chemischen Gesellschaft 31, 2958 (1898). 
* Fehling's solution heated by itself should not give a predpitate of cuprous 
oxide. 



r 



NON-VOLATILE POISONS 63 

3. Oxidation Test. — Picrotoxin, treated with a little con- 
centrated sulphuric acid in a porcelain dish, first becomes 
orange-red and then dissolves when stirred forming a reddish 
yellow solution. A drop of potassium dichromate solution will 
produce a red-brown color around the margin of the drop. If 
the two liquids are thoroughly mixed, there is an immediate 
dirty brown color wiiich passes into green on long standing. 

A green color alone is without significance, since many organic substances 
capable of reducing chromic acid to chromic oiide produce the same result. 

4. H. Melzer's' Test.- — Put some picrotoxin upon a watch 
glass and add i or 2 drops of a mixture of benzaldebyde and 
absolute alcohol. Careful addition of a drop of concentrated 
sulphuric acid will produce a distinct red color. If the watch 
glass is tilted, red streaks will run from the substance through 
the liquid. 

XTk a, freshly prepared, 30 per cent, solution of benz^ildehydc in absolute alco- 
hol. Beosaldebyde alone gives a. yellowish brown color with concentrated 
sulphuric add. Alcohol is added as a diluent to diminish this color as much as 
possible. Under these conditions the solution has a light yellow color, and the 
dark ted tint caused by picrotoxin is very clearly defined. This red color is 
unstable and, beginning at the margin, gradually fades into a pale pink or violet. 
H. Ereis' has found that cholestcrine and phytostetine' give simiUr colors with 
Melzer's reagent. 

5. Langley's Test. — Mix picrotoxin with about 3 times 
the quantity of potassium nitrate, and moisten the mixture 
with the smallest possible quantity of concentrated sulphuric 
acid. Then add strong sodium hydroxide solution in excess 
and an intense red color will appear. 

Detection of Picrotoxin in Beer 

First, neutralize the beer «ith magnesium o.ude. Then evaporate 500 cc. or 
ore to a syrup upon the water-balb. Digest this residue with 4 or 3 times its 
volume of alcohol and evaporate the alcoholic eitract. Dissolve the residue in 
hot water and filter the solution through a moistened paper. Acidify the filtrate 

' Zeitschrift fUr analytische Chemie 37, 351 and 747 {1898). 

* Chemiket-Zeitung 33, 11 (iSgg). 

• A substance very similar to cholesterine, and named paracholesterine or 
pbytosterine, is found in the seeds of certain plants. (Perkin and Kipping, 

LOrganic Chemistry, page 6o8.) 



64 DETECTION OF POISONS 

with dilute sulphuric add and extract repeatedly with ether, or better with chlo- 
roform. Evaporate these extracts and test the residue for picrotoxin. 

Should the residue from the ether or chloroform be too impure, dissolve it again 
in hot water, filter, evaporate and extract with ether or chloroform. To pvtxify 
picrotoxin further, precipitate colored substances from its aqueous sc^ution with 
lead acetate, filter and remove lead from the filtrate by hydrogen sulphide. The 
filtrate from lead sulphide upon evaporation, or extraction with ether or chl<»o- 
form, will give nearly pure picrotoxin. The very bitter taste of picrotoxin as 
well as its strong tendency to oystallize are additional characteristics of this 
substance. 

COLCmCIN 

Colchidn, CttHstNO«, an alkaloid occurring in all parts of the meadow saffron, 
Colchicum autumnale, b a yellowish, amorphous powder which is poisonous and 
very bitter to the taste. It is freely soluble in water, alcohol, and chloroform, less 
so in either, and benzene, and almost insoluble in petroleum ether. Solutions of 
colchicin have a more or less yeUowish color which becomes more pronounced 
upon addition of acids or alkalies. These solutions have very faint basic proper- 
ties. Consequently ether or chloroform, but not benzene nor petroleum ethtt, will 
extract colchidn from an add, aqueous solution. Upon evaporation of the sol- 
vent, colchicin will appear as a yellowish, sticky residue resembling a resin or 
varnish. Heated with water containing sulphuric add, colchidn splits into col- 
chicdn and methyl alcohol. Boiling the alkaloid 1.5-2 hours with 60 parts of x 
per cent, hydrochloric add will produce the same result: 

CmH„NO, -f H,0 = C,iH„NO, + CH,.OH 
Colchicin Colchicein Methyl Alcohol 

On the other hand, colchidn is formed when colchicein is heated to xoo^ with 
sodium methylate (CHt.ONa) and methyl iodide (CHt.I). Since colchicein on 
treatment with hydriodic add >'ields tEree molecules of methyl iodide, odchicein 
as weU as colchicin contains three methoxyl groups. Heated with strong hydro- 
chloric add, colchicdn loses acetic add and passes into trimethykccdchicinic 
add. Consequently colchicein and coldiidn contain an acetyl group (CH».CO — ). 
The formula of colchidn, that is to say, of methyl-colchicein, may be written as 
follows: 

CH,0\ /XH.CO.CH, 

CH,0-Ci»H,< 

CH,0/ ^CO.OCH, 

Detection of Colchicin 

Aqueous colchicin solutions, especially in presence of dilute 
mineral acids, have a yellow color. Unless the ether residue has 
this characteristic, colchicin is absent. 

I. Tannic Acid Test— This reagent will precipitate colchicin 
from aqueous solution » if not too dilute, as white flocks. This 
test, however, is not characteristic of colchicin. 



NON-VOLATILE POISONS 



65 



2. Nitric Acid Test. — Nitric acid (Sp. gr. 1.4 = 66 per cent.) 
dissolves colchicin with a dirty violet color which soon changes, 
when stirred, to brownish red and finally to yellow. Addition of 
dilute sodium or potassium hydroxide solution, until the reaction 
is alkaline, produces an orange-yellow or orange-red color. 

3. Sulphuric Acid Test.— Concentrated sulphuric add dis- 
solves colchicin with an intense yellow color. A drop of nitric 
acid added to such a solution produces a green, blue, violet and 
finally a pale yellow tone. Excess of potassium hydroxide solu- 
tion will now bring out an orange-red color, Erdmann's 
reagent (see page 313) dissolves colchicin with a blue to violet 
color. 

4. Hydrochloric Acid Test^ — Concentrated hydrochloric acid 
dissolves colchicin with an intense yellow color. Add two drops 
of ferric chloride solution and heat the mixture 2-3 minutes in a 
test-tube. The color deepens and the solution on cooling, 
especially if diluted with the same volume of water, becomes 
green or olive-green. Finally shake the solution with a few 
drops of chloroform. This solvent becomes yellowish brown, or 
garnet-red, and the aqueous solution retains its green color. 
Zeisel's reaction. 

Purificatioa of the Residue Containing Colchicin 
To isolate as pure colchicin as possible from the yellow 
residue, extract with warm water. Filter the soluUon and, 
when cold, extract it first with petroleum ether. This will 
remove fatty, resinous and colored impurities but not colchicin. 
Then extract with chloroform. Or precipitate colchicin from 
aqueous solution, which must not be too dilute, with tannic 
acid. Collect this precipitate upon a filter and wash with cold 
water. Mix the moist precipitate with freshly precipitated, 
washed lead hydroxide. Dry the mixture, grind to a powder 
and extract with chloroform. Evaporation of the solvent will 
Jieave nearly pure colchicin. 



PICRIC ACID 



oitrophenol, crysUlliies from wa 
n lemon-yellow, rhombic prisms. 



1 light yeUoi 



66 DETECTION OF POISONS 



Though soluble in cold water with difficulty, picric add dissolves freely in hot 

water, as well as in alcohol, ether and benzene. Aqueous solutions have an add 

QTT reaction, a very bitter taste and dye animal fibers fast ydlow. 

i Materia] containing picric add has a yellow or yellowish 

green color. 
r^\i n ^ysiological Action and EUminAtion.— Picric add is 
OjN Y ^ ^^« quite an active poison. Taken internally it produces a 
HC CH striking yellow pigmentation first of the conjunctiva and 
%/ then of the entire skin, usiiaUy designated as "picric add 

Y icterus.'' Picric add and its salts like most nitro-oMn- 

NO pounds decompose the red blood-corpusdes forming met- 

haemoglobin. Consequently it is a blood-poison. At the 
same time it irritates the central nervous system and causes convulsions. 
Finally it exercises its power of predpitating proteins in add solution. This is 
espedally noticeable in those organs of the body, for example, the stomach and 
OH kidneys, which, owing to necrotic tissue changes, have an 

I add or only a faintly alkaline reaction. The organism re- 

^. duces picric to picraminic add which does not so readily 

Q xj__^ Q jQ^jj predpitate protein. By thus changing picric add the 

ill organism rids itself of the poison. In picric add poisoning 

CH the urine has a marked red color owing to formation of 
^^ picraminic add. Some picric add passes into the urine 

unchanged. Elimination is slow. In one case * (see R. 



NOi Kobert, *'Intoxikationen"), after administration of a single 

Picraminic acid ^o^e of I gram of picric add, its presence in the urine coidd 

be recognized for 6 days. The urine was ruby red, dear, add and free from 

albumin and bile-constituents. Picric add was also easily detected in the feces. 

Detection of Picric Add 

Material containing picric acid has a more or less yellow or 
yellowish green color. Aqueous, alcoholic and ethereal solu- 
tions show the same color. Finely divided animal material 
should be extracted several hours under a return-condenser with 
alcohol containing hydrochloric acid to decompose compounds of 
picric add with albumins and thus bring the acid into solution. 
Filter and evaporate such an alcoholic extract upon the water- 
bath. Treat the residue, which is yeUow, yeUowish green, or 
frequently yellowish red or reddish brown, with warm water and 
filter the extract. The filtrate itself may be tested directly 
for picric acid, or it may first be extracted as usual with con- 
siderable ether. The following tests may then be applied.to the 
residue left on evaporating the ether extract: 



NON-VOLATILE POISONS 67 

z. Isopuipuric Add Test — Gently heat (50-60^) an aqueous 
solution of picric add with a few drops of saturated, aqueous 
potassium cyanide solution (1:2). The solution will become red 
owing to formation of potassium isopurpurate. One milligram 
of picric add, dissolved in 5 cc. of water, will give a distinct test, 

Isopurpuric add does not exist in the free state but is present in this test as 
the potassium salt. Nietzki and Petri^ regard isopurpuric add (CiHsOcNi) 
as a dicyano-picraminic add «■ 5-^zy-6-amino-2,4-dinitro-isophthalic nitrfle; 
whereas Borsdie' considers it a dicyano-dinitro-ozy-/3-phenyl hydrozylamine: 

OH OH 

0,N— C C— NH, OiN—C C--NH.OH 

NC— C C— CN NC— C C— CN 

Y Y 

NOf NO, 

Nietsld-Petri Borsche 

» 

2. Picraminic Acid Test — (a) Heat picric add solution with 
a few drops of sodium hydroxide solution and glucose. Picra- 
minic add, formed by reduction of picric add, colors the solu- 
tion deep red. Avoid excess of sodium hydroxide solution, 
otherwise there will be a red color due solely to the action of the 
alkali upon glucose. 

(j8) The test may also be made by wanning picric add 
solution with a few drops of sodium hydroxide and ammonium 
sulphide solutions. This will reduce picric add and produce a 
red color. 

In both reactions (a and 0) picric add is reduced to picraminic 
add, 2-amino-4,6-dinitro-phenol: 

OH OH 

OJN— C, iC— NO, 0,N— C, ,C— NH, 

Hi <!h ^^^= ni L ■*-"«'°- 
c c 

I I 

NOi NOi 

Picric acid Picraminic acid 

t Beridite der Deutschen chemischen Gesellschaft 33, 1788 (1900). 
^Ibid.« 33» 97x8 and 2995 (1900). 



68 DETECTION OF POISONS 

The presence of fat and other impurities materially influences 
this test. 

3. Dyeing Test — ^Dissolve the substance containing picric 
acid in hot water and put white threads of wool, silk and cotton 
in the solution. In a few hours (12 to 24) remove the threads 
and thoroughly rinse in pure water. K picric acid is present, 
the wool and silk will be dyed yellow but not the cotton. In 
other words, picric add is not fast upon vegetable fibers like 
cotton. Picric add, diluted i : 100,000, will still produce a yd- 
low color upon wool. 

4. Ammoniacal Copper Test — ^Add a few drops of ammonia- 
cal copper sulphate solution (copper sulphate solution and an 
excess of ammonia) to an aqueous picric add solution. A yd- 
lowish green predpitate, consisting of hexagonal needles with a 
polarizing action upon light, will appear. Picric add, diluted 
1 : 80,000, will give this test. 

ACETANIUDE 

j^jjj QQ ^jj Acetanilide crystallizes in colorless and inodorous, shin- 

I ing leaflets. It has a faint, burning taste; melts at 113 to 

C 1 14^; is soluble in 230 parts of cold water, in about 32 parts 

xy^^u of boiling water and in 3.5 parts of alcohol; and is freely 

I II soluble in ether and still more so in chloroform. AU 

HC CH acetanilide solutions are neutral. Heated to boiling with 

%/" potassium hydroxide solution (I) and also with fuming 

^ hydrochloric add (II), acetanilide is decomposed into its 

constituents: 

I. CeH».NH.CO.CH, -f KOH = C«H».XH, + CH,.CO.OK. 

II. C«H».NH.CO.CH, + HCl + H,0 = CeH».XHi.HCl + CH,.COOH. 

Physiological Action. — Being an aniline derivative, acetanilide has the 
poisonous properties of that amine though in less degree. R. Robert ("In- 
toxikationen"; refers to several instances of acetanilide poisoning which did 
not terminate fatally. In one case a student took a teaspoonful of the drug. 
There was stupor, uneasiness, marked cyanosis and lowering of the pulse. 
A purgative and restorative (stimulant) were used but there was considerable 
exhaustion for several days. The picture was nearly the same in the case of 
a man who took 2 grams of antifebrine daily for a days in succession. 

Preparation.— Boil aniline and glacial acetic acid together for several hours 
under a return-condenser: 

C.H,-NH, -f CHrCOOH - C JIi-NH-CO<:H, + H«0. 



NON-VOLATILE POISONS 69 

Detection of Acetanilide 

Ether or chloroform will extract acetanilide completely from 
an add aqueous solution. 

1. Indophenol Test. — ^Boil acetanilide with about 4 cc. of 
fuming hydrochloric add and evaporate to a few drops (about 
10). Cool and add 4 cc. of saturated, aqueous carbolic add 
solution. A few drops of caldum hypochlorite solution will 
produce a violet-red color. In time the color will become deeper, 
espedally if the mixture is shaken. Then carefully add ammon- 
ium hydroxide solution as a surface-layer which will take on a 
permanent indigo-blue color. 

The indigo-blue color is characteristic of acetam'lide only 
when preceded by the red-violet color, since a mixture of aque- 
ous phenol and hypochlorite solution gives a blue color with 
ammonia (see carbolic add). 

Phenacetine also gives the indophenol test. 

2. Phenylisocyanide Test. — ^Boil acetanilide with 5-6 cc. 
of alcoholic potassium hydroxide solution. Cool, add 2 or 3 
drops of choloroform and again heat. The offensive odor of 
phenylisocyanide will be developed. 

Potassium hydroxide decomposes acetanilide into aniline 
and potassium acetate (see Reaction I above). The former 
with chloroform gives phenyKsocyanide. 

3. Calcium Hypochlorite Test. — ^Boil acetanilide a few min- 
utes with alcoholic potassium hydroxide solution as in test 2. 
Dilute with water and extract aniline with ether. This sol- 
vent upon evaporation will deposit aniline as an oily liquid. 
Dissolve the latter in water and test with caldum hypochlorite. 



ination of Acetanilide Urine^ 

Scarcely more than traces of unaltered acetanilide appear in urine even after 
large doses. The most essential change occurring in the body is oxidation of the 
benzene ring which produces aceto-para-aminophenol. This Uke most phenols 
forms a conjugate sulphuric add and appears in the urine as a salt of aceto-para- 
aminophenyl sulphuric add: 

^ To study the behavior of acetanilide in the body, take at night 0.3 gram of 
this substance at a dose twice in the course of 3 hours and examine the urine 
passed in the next 12 hours. 



70 DETECTION OP POISONS 

H OH Q-SOr-OH 

HC CH HC CH „« HC CH 

I II +0- T I + %so«- I i 

HC CH oxidation HC CH xxt\/ HC CH 

V V V 

C C oonjogation C 

NH.CO.CH, NH.CO.CH, NH.CO.CH« 

AceUmilide Aceto-p-aminophenol Aoeto-p-aminophsii]^ aulphttric mdd 

To some extent also, a conjugate glycuronic add of aoeto-para-aminophenol is 
formed. These compounds, heated with concentrated hjrdrocfaloiic acid, give 
para-aminophenol which can be detected by the indophenol test previously 
described. 

O.SOt.OH OH 

i A 

HC CH HC CH 

I II + 2H,0 = HtS04 + CH,.COOH + I jl 

HC CH HC CH 

Y y 

I I 

NH.CO.CH, NHt 

p-«minophenol 

Such urine, boiled a few minutes with concentrated hydrochloric add, will 
usually give the indophenol test. But the test will be more certain, if para« 
aminophenol is first isolated. Boil a larger quantity of urine (300 to 500 cc.) a 
few minutes with about 10 cc. of concentrated hydrochloric add. Then add an 
excess of sodium carbonate and repeatedly extract the cool urine with large 
quantities of ether. Distil or evaporate the ether. Para-aminophenol usually 
appears as a reddish or brownish oil. An aqueous solution of this substance will 
give the indophenol test. 

PHENACETINB 

Phenacetine, or p-aceto-phenetidine, oystallizes in shining leaflets, which are 

without color, odor or taste, and mdts at 134 to 135^. Phenacetine is soluble in 

^TT pQ prj about 1400 parts of cold water, 70 parts of boiling water, 

I 16 parts of alcohol and fredy soluble in ether and 

C chloroform. Its solutions are neutraL Concentrated sul- 

^\ phuric add dissolves it without color. Phenacetine is 

HC CH 

I jT very dosely related to acetanilide but does not give the 

jjC CH phenylisocyanide test. 

%/ Preparation. — The gradual addition of crystallized 

^ phenol to cold dilute nitric add (9. gr. i.ii » X7.5 per 

Ip TT cent.) results in the formation of a mixture of o- and 

p-nitro-phenol. Since the ortho-compound is volatile with 
steam, complete separation of the two products is possible by steam distilla- 
tion. The residual p-nitro-phenol is converted into its aodiom salt whidi ii 



NON-VOLATILE POISONS 



71 



heated in sealed tube with eth^ bromide and thus changed to p-nitro-phenetoL 
The latter is reduced by means of nascent hydrogen from tin and hydrochloric 



acetic add and converted into w 


ceto-p 
ONa 


-phenetidine, 


or phenacetine: 


OH 


iOH H 


0;Na" 


Bri.CHi oCHi 


i 


V 




c 




i 


HC CH 


HC CH 




HC CH 




HC CH 


HC CH ~* 


ni L 


— ♦ 


T 1 

HC CH 


-♦ 


HC CH 


C 


Y 




Y 




Y 


\ HO —NO, 


NO, 




1 

NO, 




n[o, 1 

a Hi 4H| 


Phenol 


p-NHro- 




Na salt of p- 




p-Nitro^" 




phenol 




nitro-phenol 




phenetol 




OC,H, 






OC,H, 






} 






i 






HC CH 




H 


C CH 






Hi k 




— > 

H 


i k 






Y 




• 


Y. 






HN;H i 






NH.CO.CH, 




|HO;.CO.CH, 










p-Phenetidine 






Phenacetine 



Detection of Phenacetine 

The extraction of phenacetine by ether or chloroform from 
an aqueous tartaric add solution is complete. 

1. Qzidatioii Test — Boil phenacetine for several minutes 
with 3 cc. of concentrated hydrochloric acid. Dilute with 
lo cc. of water and filter when cold. A few drops of chromic 
add solution added to the filtrate will gradually produce a 
ruby red color. Strong chlorine water may be substituted 
for chromic add. 

2. Indophenol Test. — Boil phenacetine i or 2 minutes with 
about 2 cc. of concentrated hydrochloric add. Dilute with 
water and add a few cc. of aqueous carbolic add solution. 
Filter the solution when cold. If a few drops of freshly pre- 
pared caldum hypochlorite solution are added, the filtrate will 



72 DETECTION OF POISONS 

have a fine carmine red color. Addition of ammonium hydrox- 
ide solution in excess will change this color to violet-blue. 
Freshly prepared chlorine water, or 3 per cent, chromic add 
solution, may be substituted for hypochlorite solution as an 
oxidizing agent. 

3. Autenrieth-HinsbergTest.^ — (a) With Dilute Nitric Add. 
Heat phenacetine to boiling with a few cc. of dilute nitric 
acid (10 to 12 per cent.). It is soluble and gives an intense 
yellow to orange-red color. As the solution cools, if sufficiently 
concentrated, nitro-phenacetine^ will aystallize in long, yellow 
needles which melt at 103°. This test is delicate, and char- 
acteristic of phenacetine, especially when nitro-phenacetine can 
be obtained in crystals and its melting point determined. It 
serves to distinguish phenacetine from acetanilide and anti- 
pyrine, both of which give colorless solutions when warmed with 
dilute m'tric acid. 

(b) With Concentrated Nitric Acid. — ^A few drops of con- 
centrated nitric acid poured upon phenacetine produce a yellow 
to orange-red color. Part of the phenacetine is dissolved with 
the same color and heat completes the solution. Nitro-phen- 
acetine crystallizes as the solution cools. 



H 



SALICYLIC ACID 

COOH Salicylic add, or ortho-oxy-benzoic add, crystallizes in long, 

I white needles soluble in about 500 parts of cold and in 15 

C parts of boiling water; and freely soluble in alcohol, ether, 

-ar r nw chloro^orni and carbon disulphide. It has a peculiar taste 

I II which is sweetish, addulous and rather acrid. It melts at 157^. 

HC CH Heated carefully, salicylic add will sublime in fine needles 

"S^ without decomposition. A little of the add may show this 

behavior even upon the water-bath. If heated quickly, salic^c 
add is decomposed in part into phenol and carbon dioxide. 
yCOOH 

C.H4 = CeHj-OH + CO,. 

^ Archiv der Pharmade 229, 456 (1891). 

' The structural formtila of mono-nitro-phenacetine is as follows: 

CHr-NO, 3, 

\nh(c,h,o) 4 



NON-VOLATILE POISONS 73 

Concentrated sulphuric add dissolves pure salicylic add without color and 

without decomposition. The lead and silver salts of this add are soluble in water 

with difficulty. Consequently lead acetate will predpitate lead salicylate, 
.COOn 



( C.H4 ) Pb, 



from neutral solutions. This salt is white, oystalline and 

OH 

soluble in hot water. It crystallizes unchanged as the hot solution cools. Silver 
nitrate predpitates white silver salicylate. 

IL Sdimitt's Method of Preparation. — Dry sodium phenolate is kept cool and 
saturated in an autoclave under pressure with carbon dioxide (a). The sodium 
phenyl-carbonate undergoes molecular rearrangement, when heated at 120-130% 
and becomes isomeric sodium salicylate (/3) : 

^O ONa /ONa ^^^^ /OH (i) 

(a) C 4- I =» C=0 OS) rearrangement CeH4 

V) CeH, \)C.H, 8^^^* \cOONa (2) 

Sodium phenyl- Sodium salicyl- 

carbonate ate 

Detection of Salicylic Acid 

1. Ferric Chloride Test. — Addition of ferric chloride solution 
to a solution of salicylic acid or salicylates produces a blue-violet 
color. If the solution is very dilute, the color is more of a red- 
violet. Hydrochloric acid changes the violet color to yellow. 
An excess of the reagent aflfects the delicacy of the test. 

This test fails in presence of mineral adds, caustic alkalies and alkaline 
carbonates. 

2. Millon's Test. — ^If an aqueous salicylic add solution is 
warmed with Millon's reagent, a deep red color will appear. 

3. Bromine Water Test. — This reagent in excess produces a 
yellowish white, crystalline precipitate even with very dilute 
salicylic add solutions. The compound thus formed is tri- 
bromo-phenyl hypobromite (see page 28). 

OBr 

<! 

OH ^^ 

/"" BrC CBr 



C.H4 + 4Brt = COj +4HBr + I 

C 
Br 

4. Melting-Point Test — If the quantity of salicylic add is 



74 DETECTION OF POISONS 

not too small, dissolve the ether residue in very little hot water, 
shake the hot solution with a little animal charcoal and filter. 
Cool the filtrate, dry the crystals and determine the melting 
point (iS7°)- 

Separation of Salicylic Add'from Simple Phenols 

When phenols like carbolic acid or the cresols are present, 
the above tests from i to 3 prove nothing as far as salicylic 
add is concerned. If these compounds are present, add suffi- 
cient sodium carbonate solution to render the ether residue 
alkaline and extract the solution with ether. This solvent 
will take up the phenols and salicylic acid will remain in the 
water as the sodium salt. Withdraw the aqueous solution 
from the separating funnel, acidify with dilute hydrochloric 
or sulphuric acid and extract salicylic acid with ether. 

Directions are given elsewhere (see page 243) for the detection 
of salicylic acid in beer, milk, urine, friiit juices, meat and meat 
preparations, as well as in maltol. 

Quantitative Estimation of Salicylic Add as Tribromo-phenyl Hypo- 

bromite 

Place the aqueous solution of saUcylic acid in a glass-stop- 
pered flask, add an excess of saturated bromine water and shake. 
The acid is completely precipitated as tribromo-phenyl hypo- 
bromite. At the end the solution should be reddish brown. 
It should stand 12-24 hours and be shaken frequently. Collect 
the precipitate of tribromo-phenyl hypobromite in a weighed 
Gooch crucible and dry to constant weight in a vacuum desic- 
cator over sulphuric acid. The quantity of salicylic add may 
be calculated from the weight of precipitate as follows: 

CeHiBr*©: C7HeOi = Wt. of precipitate : x 
409.86 138.05 obtained 

Detection of Salicylic Add in Urine^ 

Salicylic aad forms a conjugate with glycocoll, supplied by the organism, 
and is changed in the human body, in part at least, into salicyluric add, 

* To study the behavior of salicylic acid in this connection, take i to 1.5 grams 
of sodium salicylate at night in the course of several hours and examine, as de- 
scribed, the urine passed in the next 1 2 hours. 



NON-VOLATILE POISONS 76 

(i) 



X:0.NH.CH,J 



C,H4_ 

.COOH (2) 

Thb is eUminated in urine with unaltered salicylic add. Such urine gives a 
violet color with ferric chloride solution. Both salicylic and salicyluric adds 
give this test. To decompose salicyluric add into its constituents, heat the add 
half an hour with fuming hydrochloric add under a return-condenser. 

To isolate unchanged salicylic add, addify 500 to 1000 cc. of urine with hydro- 
chloric add and repeatedly extract with ether. Remove the ether from the 
aqueous solution in a separating funnd and shake vigorously with excess of 
sodium carbonate solution. Salicylic add passes into the aqueous solution. 
Withdraw the aqueous solution, which is alkaline, addify with dilute hydro- 
chloric add and extract with ether which upon evaporation usually deposits 
the add in a crystalline condition. Purify the residue by recrystallization from 
water, using animal charcoal to remove color. Salicylic add is rapidly taken up 
by all mucous surfaces and quickly absorbed. Elimination by way of the urine 
usually begins within the first half hour and is complete in 3 da3rs. 

VERONAL 

Veronal, C-diethyl-barbituric add, C-diethyl-malonyl-urea, CtHiaOiNs, 
crystallizes from hot water in large, colorless, spear-shaped crystals melting at 
CjHrv /CO — NHv 191** (corrected). It is soluble in 146-147 parts of 

/^\ yCO water at 20® and in 15 parts at loo*. Veronal is also 

CiHf CO NH freely soluble in hot alcohol and in acetone. It dis- 

solves with difficulty in cold ether. An aqueous veronal solution has a bitter 
taste and shows a very faint add reaction with sensitive blue litmus paper. 
Veronal readily dissolves in caustic alkalies, ammonia and in caldum or barium 
hydroxide solution. From such solutions, provided they are not too dilute, 
adds repredpitate veronal in a crystalline condition. Of the veronal salts the 
sodium salt, CtHnOiNsNa, crystallizes best. It may be prepared by dissolving 
veronal in the calculated quantity of caustic soda solution free from carbonate, 
and then evaporating this solution with exclusion of carbon dioxide, or adding 
alcohol until turbidity appears. In both cases the sodium salt of veronal sepa- 
rates as splendid shining crystals. 

Preparation by £. Fischer and A. Dilthey^ 

(a) From diethyl-ethylmalonate by condensation with urea in presence of 
sodium ethylate: 

NajOCaH* 
CiHi CO— ;OCSU HN iH C,H» CO— N— Na 

)>C<( ^^ ^ y^^ y^^ ^ 3C,He.0H. 

CiHi CO— !0C,H6 HNH CH* CO— NH 

Diethyl-ethylmaionate Urea Na salt of veronal 

* Annalen der Chemie und Pharmazie, 335, 334 (1904). 



76 DETECTION OF POISONS 

Dissolve metallic sodium (32 parts) in absolute alcohol (600 parts) and when 
cold add diethyl-ethylmalonate (100 parts). Dissolve in this mixture with heat 
finely powdered urea (40 parts). Heat 4-5 hours in an autoclave at 105-108^. 
The sodium salt of veronal is precipitated even from the hot solution as a color- 
less, crystalline mass. Cool, filter with suction and wash with alcohol. Dis- 
solve the crystals in water and acidify with concentrated hydrochloric add. 
Veronal thus precipitated is pure when recrystallized from water. 

(b) From diethyl-malonyl chloride by condensation with urea: 

C,H, CO— ;Ci H:NH C,H, CO— NH 

Nc/ NCO = 2HCI + y^^ /^CO 

CjHi CO— ICl H;NH C,H, CO— NH 

Heat diethyl-malonyl chloride (3 parts) on the water-bath for 20 hours with 
finely powdered, dry urea (2 parts). Considerable hydrochloric add is given 
ofif toward the end and a solid mass finally remains which yields pure veronal 
upon crystallization from hot water. The yield is 70 per cent of the theoretical 
amotmt. 

Physiological Action. — Veronal does not cause decomposition of the blood and 
in the usual medicinal doses (0.5-1 gram) does not appear to act strongly upon 
the heart. Cumidative action has been noted only in rare instances. In large 
doses, however, veronal may cause serious intoxication with fatal termination. 
Death resulted in the case of a man in Holzminden who had taken 10 grams of 
veronal by accident. There are also on record two other suicidal cases, one from 
II grams and the other from 15 grams of this hypnotic. There was loss of con- 
sdousness and contraction of the pupils in the second case. Atropine caused 
dilatation of the pupils but otherwise was of no avail. Death ensued in 20 hours. 



Detection of Veronal 

In examining cadaveric material (liver, spleen and kidneys) 
from a man, who had taken veronal, thinking it was a remedy 
for tape- worm (kamala), G. and H. Frerichs^ isolated small 
quantities of this drug. Following the Stas-Otto process, they 
extracted the aqueous tartaric acid solution with ether and 
evaporated the ether extract. They recrystallized the residue 
from a little hot water, using animal charcoal to remove color, 
and identified the crystals obtained as veronal by the following 
tests: 

1. The aqueous solution of the crystals had a faintly add 
reaction. 

2. The crystals were soluble in sodium hydroxide solution 

^ Archiv der Pharmazie 244, 86-90 (1906). 



NON-VOLATILE POISONS 77 

and ammonia and were reprecipitated when the solutions were 
acidified with dilute hydrochloric acid. 

3. The melting-point of the crystals was 187-188°. On 
mixing the substance with pure veronal, they obtained the same 
melting-point. ^ 

4. Presence of nitrogen was shown by fusing the crystals in a 
dry test-tube with metallic sodium, cooling the melt, dissolving 
in water and testing for sodixim cyanide by the Prussian blue 
reaction (see page 2 2) . 

5. The crystals were heated in a dry test-tube and sublimed. 
They were then compared with crystals known to be pure 
veronal and found identical. 

Detection of Veronal in Urine 

E. Fischer and J. v. Mering,^ and also B. Molle and H. EUeist,' 
have found that most^of the veronal leaves the human body 
unchanged and is present in the urine to the extent of 70-90 
per cent. Consequently in veronal poisoning the urine should 
be examined first. Concentrate a considerable quantity on 
the water-bath* to one-fifth its volume and extract several 
times with ether, using a large volume at each extraction because 
veronal is not very soluble in this solvent. The residue left 
after distilling the ether is usually quite dark in color. Dissolve 
in as little hot water as possible, boil the solution 15 minutes 
with animal charcoal and filter. Cool the nearly colorless 
filtrate with ice and veronal will crystallize in colorless needles 
melting at 191° (corrected). 

£. Fischer and v. Mering recovered from 5 days urine, after administration 
of 4 grams of veronal during 2 days, 2.49 grams » 62 per cent, of the quantity 
used. The method therefore is not absolutely quantitative nor is the elimination 

^ A mixture of two organic substances, having the same melting-point but not 
being identical, will show a melting-point lower than that of either substance 
taken by itself. But obviously a mixture of identical substances will show no 
depression of melting-point whatever. 

* Die Therapie der Gegenwart 45, 1904. 

' Archiv der Pharmazie 242, 401 (1904). 

^ Fischer and v. Mering evaporated the urine under diminished pressure. 



78 DETECTION OF POISONS 

of veronal . complete after 5 dajrs. The ciystals obtained should be proved 
conclusively to be veronal by the tests described. 

Molle and Kleist first add lead acetate solution to the urine as long as it causes 
a precipitate, filter, remove lead from the filtrate by hydrogen sulphide and filter 
from lead sulphide. Hydrogen sulphide is expelled with heat and the urine, 
after dilution with twice its volume of water, is boiled with animal charcoal. 
The filtrate, after concentration on the water-bath to a small volume, is cooled, 
saturated with sodium chloride and extracted 3 times with ether. The 
filtered ether solution on distillation leaves nearly pure veronaL 

AMTlPyJUMK 

Antipyrine, or i-phenyl-3,3-dimethyl-isopyraa<done, CuHnONi, forms 

monocJinic, tabular cr>-stals having a faintly bitter taste and melting at 1x3®. 

, \ «>tx f^ x^ #^u / \ ^c P&rt of antip>*rine is scduble in less than x 

) I part of cold water, m about x part of alcohol, x 

HC Sr — C«H« (i) part of chloroform and in about 50 parts of 

\. ether. .\n aqueous antipyrine scdution has a 

|t neutral reaction, although this compound is a 

base and forms cr>-sial1i«able salts with adds. 
Preparatioii. .\ntipyrine b formed directly by heating ^-phensi-methyl- 
h\*drajdne and avxtosict^tic ester: 

CH,— C O H X— CH, V.0 CHi— C — X— CH, (2) 

HC^ H H X-C»H» - HC X— C«BU (i) + H^ + C,H».OH. 

X , O C»H» C 

Dettctkn of Antipjiine 

Kthor c:ttrnctii only :im)all quantities of antq>yrine from a 
solution \\M)tauuni; much tartaric acid. Ether, or better 
chlorvMorm. extracts by lar the greater part of the antq>>Tiiie 
when the solution has In^n made alkaline. Antqjvrine diffars 
CrvMu nuxst alkaUxi\ls in Wing mort^ svxhihle in water. To detect 
antipvrinc. dissolve the residue left on evaporating the ether 
solution in a little wAter. lUter and apv^ly the following tests: 

I. F^MTic Chlc4ridi> TVat. Add i v>r -* drv^x? of f«ric dikdde 
svxlution to an a\)u«vus anti\^Yiine s\>lution. It will pfodnce 
a d^vp ivnI kwUw \\l\ich van Iv seen ex-t^n ia a dihiticai of i : 

i. T^umic Acid T^t. Tannic aciv: svVutioD prvxhices an 
abundant, white pusijxitato. x^hen avWev: :o an aqueous anti- 
pyrine solutivMX 



NON-VOLATILE POISONS 



79 



3. Fuming Nitric Acid Test. — Dissolve antipyrine in a few 
drops of water and add 1 or 2 drops of fuming nitric add. The 
solution will be green. If this solution is heated to boiling, 
another drop of nitirc acid will produce a red color. Two cc. 
of antipyrine solution (1 : 200) will give this test distinctly. 

4. Nitroso-antipyrine Test. — Add a few drops of potassium 
or sodium nitrite solution to an aqueous antipyrine solution 
and then dilute sulphuric acid. A green or blue color will 
appear. A few drops of acetic acid may be substituted for 
sulphuric acid but the solution must be heated. If the anti- 
pyrine solution has been concentrated, green crystals of nitroso- 
antipyrine (CiiHii(NO)ONi) will separate after some time. 



Detection of AntipTrine in Uiine 

Part of the antipyrine passes unchanged into the urine but some is also present 
as oxy-anlipyrine-glycuronic acid, a direct test (or which may be made by means 
of ferric chloride solution. When the urine is highly colored, render a consider- 
able quantity alkaline with sodium hydroxide solution or ammonia and Extract 
with chloroform. Dissolve the residue left by chloroform in a little water nod 
test for antipyrine with ferric chloride solution and with fuming nitric add. 



with ehlo: 
test for ai 

ICaffeini 
oyatalliu 
(I) CHr- 
(.) . 
43) CHr- 
fn ether. 



CAFFEINE 

Caffeine (theine) or i.3,7-trimelhy!-j,6-dioiy-purine {C,HnO,Ni.H,0). 
oyatalliies in while, shining needles. It is soluble in 80 parts of water, gii'ing 

fil CH N CO(6) " colorless solution nith a neutral reaction and 

I 1 yCHi(7) a faint, hitler taste. Caffeine is quite easily 
(*) OC C— N^ soluble in hot water (1:3). Il requires for 

X J| -tj^ solulioQ nearly 50 parls of alcohol, only 9 

parts of chloroform and is only slightly soluble 
!d ether. In crystallizing from hot water caffeine combines with i molecule of 
water, a part of which it loses upon eiposure to dr and all when dried at 100°. 
Cafieine is only very slightly soluble in absolute alcohol, benzene and petroleum 
eLhet. It melts at 130', but somewhat above tod° begins to volatilize in small 
quandty and at iSo° to sublime without leaving a residue. Concentrated sul- 
phuric and nitric adds dissolve it without color. Caffeine is a very weak base 
and its salts are decomposed by water. Therefore, caffeine can be extracted at 
least partially by ether, or better by chloroform, from an aqueous tartaric add 
solution. The relation existing between caffdne and uric add is quite apparent 
when the products, formed by oadizing these two substances with potasdum 
chlorate and hydrochloric add, are compared. Oxidation of uric add yields 
alloxan and urea; caffdne gives dimethyl-alloxan and monomethyl-urca. 



80 



DETECTION OF POISONS 



CHr-N— CO 



CH, 



I I 
DC C 



— N;--CH, + (H,0) + 0,= 



\ 



CH 



Caffeine 



CHr— N— CO NH— CH, 

OC CO 4- CO 

CHr-N— CO NHt 

Dimethyl- Monomethid- 
allozan 



Fate of Caifdne in Humin Metabolism. — Only a very small part of the caffeine 
taken into the body passes through unchanged and appears in the urine. About 
lo per cent, appears in the urine as decomposition products. The remainder 
may be changed into normal end-products of human metabolism. Most of the 
nitrogen of caffeine is eliminated as urea. A very important fact is the cleavage 
of methyl groups with formation of the first decomposition products of caffeine, 
namely, dimethyl- and monomethyl-xanthines. Of the monomethyl-xanthines, 
7-monomethyl-xanthine is formed especially. Of the dimeth^-xanthines, 
parazanthine ■■ x,7-dimethyl-zanthine is fotmd. Both of these compoimds 
appear in urine after administration of caffeine. Parazanthine is isomeric 
with theophylline, or i ,3-dimethyl-zan thine, and with theobromine, or 
3,7-dimethyl-zanthine. 

The structural formuls of these cleavage-products of caffeine in animal meta- 
bolism are as follows: 



UN— CO 

r I /CH, (7) 

J: c— N< 



OC 

I 



'\ 



)CH 
HN— C— N' 
7- M et hyl •zant hine 



(i) CH,.N— CO 

OC C— NH' 



(3) CH,. 



I \\ Vh 

X— c— X^ 



Theophylline 



(x) CH,.X— CO 

OC C— X.CH, {7) 

HX-C— X 

pATAXAnthine 



HX— CO 



OC 



(3) CH,.N 



I 
C— X.CH, (7) 



Theobromine 



Detection of CalMne 

Ether will extract more catTeine from an aqueous alkaline 
solution than from an aqueous tartaric acid solution. Since 
caffeine dissolves with some difficulty in ether, but more easily 
in chloroform, the latter solvent is usually employed after the 
solution has lHH>n made alkiUine with ammonia. After dis- 
tillation of si^lvent, caffeine apjH^ars in concentric clusters erf 
long, shining neeiUes, In an analN-sis by the StasOtto method 
caffeine will apjHwr in all three extracts. 



NON-VOLATILE POISONS 



81 



I 



lation Test.- — Pour a few cc. of saturated chlorine 
water' over caffeine and evaporate the solution to dryness upon 
the water-bath. A reddish brown residue will remain. If a 
few drops of ammonium hydroxide solution are added, a fine 
purple red color will immediately appear. This test may be 
made by covering the dish containing the residue with a glass 
plate moistened with a drop of strong ammonia. Or two 
matched watch glasses may be used, the material containing 
caffeine being evaporated to dryness with chlorine water upon 
one glass which is then placed for a short time upon the other 
glass containing a drop of strong ammonia. 

This test, known as the murexide reaction, is also given by 
xanthine, theobromine, i- and 7 -monomethyl -xanthine and 
paraxanthine, especially if made as described by E. Fischer.* 
Heat the material to boiling in a test-tube with strong chlorine 
water, or with hydrochloric acid and a little potassium chlorate, 
evaporate the liquid to dryness in a dish and moisten the residue 
with ammonia. 

2. Tannic Acid Test. — This reagent, added to an aqueous 
caffeine solution, causes a heavy white precipitate which is 
soluble in an excess of the acid. This test is not characteristic 
of caffeine, 



ition of Bther Extract of Alkaline Solution 

(Mo'st of the alkaloids appear here) 

Add enough sodium hydroxide solution to the acid solution 
separated from ether to make it strongly alkaline. The alkali 
will Uberate alkaloids from their salts and combine with mor- 
phine and apomorphine, if present. Thoroughly extract this 
alkaline solution with about the same quantity of ether. 
This solvent will dissolve all alkaloids except morphine, 
apomorphine and narceine. Separate the ether from the 
aqueous solution and again extract with a fresh quantity 
of ether. In certain cases 3 or 4 such extractions may 

' A convenient method of preparing a saturated, aqueous chlorine solution is to 
heat potassium chlorate with hydrochloric add and pass the chlorine into a small 
quantity of water. 

Berichte der Dcutschen chemischen Gescllschaft 30, 2136 (1897). 



82 



DETECTION OF POISONS 



be required. Pour the ether extracts into a dry flask| stop- 
per loosely and set aside for i or 2 hours. A few drops 
of Water always settle to the bottom of the flask. Carefully 
decant the ether and pour through a dry filter. Evaporate the 
filtrate with gentle heat in a glass dish (8 to 10 cm. in diame- 
ter). Let the last part of the ether solution evaporate spon- 
taneously. If small globules having a strong odor appear, the 
residue must be examined for coniine and nicotine. If there is 
no trace of these volatile alkaloids, gently heat the residue 
upon the water-bath to expel water left by evaporation of the 
ether. Remove the dish from the water-bath as soon as this 
has been accomplished. It is not advisable to heat the residue 
too long, as it tends to become viscous. This residue, obtained 
by extracting the alkaline solution with ether, may contain 
any alkaloid except morphine, apomorphine and narceine. It 
should be examined for 



Coniine 


Brudne 


Codeine 


Quinine 


Nicotine 


Atropine 


Narcotine 


Caffeine 


Aniline 


Scopolamine 


Hydrastine 


Antipyxine 


Veratrine 


Cocaine 


Pilocarpine 


I^rramidone. 


Strychnine 


Physostigmine 







First, note the general appearance of the residue and then 
examine with the microscope. Taste it cautiously. Certain 
alkaloids may be recognized beforehand by this test. Special 
tests should then be made at once. The various alkaloids 
appear in the residue as follows: 

Strychnine. — Fine needles having an exceedingly bitter taste. 

Brucine. — Usually a white, amorphous powder having a 
very bitter taste. 

Veratrine. — Usually an amorphous powder having a sharp, 
burning taste. 

Atropine and Quinine. — A varnish which is resinous and 
sticky. Rarely crystalline or in the form of a powder. 

Codeine. — A thick, viscous syrup which after a time becomes 
solid, especially if stirred with a glass rod, and frequently 
crystalline. 



NON-VOLATILE POISONS 



83 



Caffeine.— Long, silky needles having a faintly bitter taste. 
These are frequently concentrically arranged, 

Antipyrine. — A syrup which gradually becomes crystalline, 
especially if stirred. It has a mild, bitter taste and dissolves 
very easily in water. ^ 

Pyramidone. — Usually as fine needles which have a faintly 
bitter taste. It is easily soluble in water. 

Frequently ether leaves only a slight, tasteless residue. In 
that case alkaloids are absent. Such residues often consist 
of fat, resinous matter, or traces of nitrogenous substances 
(Peptones and their cleavage products? Creatinine?). Parts 
of a cadaver, even when quite fresh, usually give small residues 
at this point. Alkaloids may be absent and every step in the 
process may have been performed with the greatest care. To 
be quite sure that alkaloids are absent, dissolve a portion of the 
residue in water containing a drop of dilute hydrochloric acid. 
Filter, if necessary, and distribute this solution upon several 
watch-glasses. Test with the following alkaloidal reagents; 



Mercuric Chloride 
lodo- Potassium Iodide 
Potassium Mercuric Iodide 
Potassium Bbmuthous Iodide 



Picric Acid 

Tannic Add 
Phospho-Molybdic Acid 
Phospho-Tungstic Acid. 

Unless these reagents give distinct and characteristic pre- 
cipitates, alkaloids are absent. It is advisable in every instance 
to make this preliminary test for alkaloids. Only a small por- 
tion of material is required and these general reagents show even 
traces of alkaloids. 

To exclude mistakes and oversights in toxicological Boalysis, dissolve the ether 
residue, should it be very small, in a few cc. of very dilute hydrochloric add (about 
I per cent, of HCl). Evaporate Ihb solution upon the water-bath and dissolve 
the residue in a little wnler. Inject this solution from a hypodermic syringe into 
the lymph-sac on the back of a smaQ but lively frog. If the frog shows no sign of 

' Most of the alkaloids are only slightly soluble in cold water. Some canoot be 
detected satisfactorily by purely chemical means. Others have no characteristic 
tests. Such substances should not be selected for laboratory practice. They 
may cause beginners to think that the experienced toxicologbt relies upon similar 
^VBCETtain methods when he seeks to identify an alkaloid in an actual analysis. 



84 DETECTION OF POISONS 

poisoning in the course of several hours, it is quite likely that the residue does not 
contain any very poisonous alkaloid. 

In making special tests for alkaloids, distribute the residue 
upon several watch glasses, using a platinum or nickel spatula 
or a small penknife. Or dissolve the residue in a little hot 
alcohol, filter the solution, distribute upon watch glasses and 
evaporate at a gentle heat. R. Mauch^ dissolves the residue 
in 75 per cent, aqueous chloral hydrate solution and uses this 
solution in testing for alkaloids. (The details of this method 
will be found on page 244.) 

Purification of the Alkaloidal Residue 

If alkaloids are contaminated with greasy, resinous or fatty 
substances, many of the tests will either fail entirely or give 
uncertain results. In this case the residue must be purified 
in one of two ways. 

1. Thoroughly mix the residue with cold water containing 
hydrochloric acid. Filter to remove insoluble matter (fatty 
or resinous substances), add sodium hydroxide solution to the 
filtrate until alkaline and extract with ether. The alkaloids 
obtained by evaporating the solvent are usually quite pure. 

2. Or dissolve the residue in hot amyl alcohol, extract this 
solution with a few cc. of very dilute sulphuric acid and with- 
draw the acid solution from the separating funnel. Amyl 
alcohol will retain greasy and colored impurities, and the alka- 
loids will be in the aqueous solution as sulphates. Add sodium 
hydroxide solution in excess and extract with ether. This 
method of purifying the alkaloidal residue is especially recom- 
mended, when there is considerable coloring matter. 

W. H. Warren and R. S. Weiss* have suggested picrolonic add* as a means of 
purifying alkaloids. An alkaloid like strychnine, whose picrolonate is very insolu- 
ble, may be precipitated from aqueous solution and thus separated from other 
substances which prevent purification. The precipitated picrolonate may be 

* Richard Mauch (Mittheilungen aus dam Institut des Harm Prof. Dr. E. 
Schaer in Strassburg), "Festgabe das Dautschen Apotheker-Vereins," Strassburg, 
1897. 

* The Journal of Biological Chemistry, 3, 330 (1907). 
' For the preparation of this reagent, see page 313. 



NON-VOLATILE POISONS 85 

collected on a filter, washed with water and then warmed with dilute sulphuric 
add which discharges the bright yellow color of the picrolonate causing the alka- 
loid to pass into solution and precipitating pale yellow picrolonic add. By 
extracting with acetic ether, in which picrolonic add is espedally soluble, the 
aqueous solution of the alkaloid is left colorless. Neutralization with sodium 
hydroxide solution and extraction with ether will give a very pure alkaloid. 

CONIINE 

Coniine, a-normal-propyl-piperidine, CgHifN, occurs in all parts of spotted 

hemlock (Conium maculatum) together with n-methyl-coniine, conhydrine, 

H^ 7-coniceine and pseudo-conhydrine. It b a color- 

C less, oily, very poisonous liquid which becomes 

/\ yellowish or brown in contact with air and is par- 

J^«9 9^* tially resinified. It is sUghtly soluble in cold but 

HjC *CH.CHs.CHj.CH| ®^^^ ^^^ soluble in hot water. Coniine is misdble 

X/ with alcohol, ether, chloroform and benzene in all 

^ proportions. The unpleasant, narcotic odor of this 

alkaloid, sometimes said to resemble that of mouse 
urine, is more intense than the odor of nicotine. Coniine as it occurs in 
nature is dextro-rotatory,^ [ajo = +18.3®, and rather a strong base. Heated 
with acetic anhydride, it forms acetyl-coniine: 

ChVcO.O.CO.CH, " C.H1.N.CO.CH, + CH,.COOH; 

Shaken with benzoyl chloride and sodium hydroxide solution, it forms benzoyl- 
coniine: 

C^CO CI + ^^^^ " C»HiJSr.CO.C»H» + NaCl + H,0; 
and with nitrous add nitroso-coniine: 

^'^iJo OH = C8HieN.N0 + H2O. 
All these reactions show that coniine is a secondary base. 



Detection of Coniine 

The alkaloidal reagents especially delicate with coniine are: 
iodo-potassium iodide (i:8ooo), phospho-molybdic add 
(1:5000), potassium mercuric iodide (1:8000) and potassium 
bismuthous iodide (1:5000). Gold and platinum chlorides 
fail to precipitate coniine when the concentration is less than 

^ The optical activity of this alkaloid is occasioned by the presence of the asym- 
metric carbon atom marked with an asterisk in the structural formula.- 



86 DETECTION OF POISONS 

I : loo; whereas they will precipitate nicotine when the concen- 
tration of the solution is as low as i: 10,000 and 1:5000. 
When coniine is present, the residue left by the ether solution 
has the characteristic odor of this alkaloid. The two following 
tests should then be made: 

1. Solubility Test — Dissolve a drop of coniine in just enough 
cold water to give a clear solution. (Jently heat the solution 
and it will become milky, because coniine is more easily soluble 
in cold than in hot water. A coniine solution which is milky 
when hot becomes clear on cooling. Aqueous coniine solutions 
have an alkaline reaction. Test the solution with red litmus 
paper. 

2. Crystallization Test. — Put a little coniine upon a watch 
glass, or glass slide, and add i or 2 drops of hydrochloric acid. 
Evaporate to dryness and coniine hydrochloride (CgHizN.HCl) 
will remain. Immediately after evaporation examine this resi- 
due with a microscope magnifying about 200 times. The color- 
less or faintly yellow crystals are needle-like, or columnar and 
frequently grouped in star-shaped clusters. They show the 
play of color characteristic of doubly refractive substances. 

NICOTINE 

Nicotine, CioHi4Na, is a colorless hygroscopic liquid which soon turns yellow 
and then brown upon exposure to air and in time becomes resinous. It is misdble 

with water in all proportions (distinction from coniine) 
9 /-TT nxj 2ind freely soluble in alcohol, ether, amyl alcohol, bcn- 
/«5^ I I zene and petroleum ether. Ether extracts nicotine 

HC ^C — CHa CH2 from aqueous solution. It has a sharp, burning taste 
y I \ / and strong odor of tobacco especially when warm. 
^^ P I Chemically pure nicotine is said to be almost inodorous. 

j^j Qn^ The so-called tobacco odor is developed after the alka- 

loid has been for some time in contact with air. The 
free alkaloid is strongly laevo-rotatory, [ajo = —161.55®, but its salts are 
dextro-rotatory. 

Constitution. — Nicotine is a rather strong di-add, ditertiary base and forms 
well-crystallized salts with one or two equivalents of add. Like ditertiary bases 
it combines with two molecules of methyl iodide* forming a di-iodo-methylate, 

1 In methyl iodide — as well as in other alkyl haloids — we have an excellent 
means of recognizing the tertiary nature of a nitrogen base. Like trimethylam- 



NON-VOLATILE POISONS 87 

» 

CioHi4Nt.3CHaI. Oxidized with chromic add, nitric add or potassium per- 
manganate, nicotine is converted into nicotinic add, or /3-carbozy-p3rridine. 
This shows that nicotine is a pyridine derivative having a side-chain in the ^- 
position with respect to the pyridine nitrogen. 

H H 

C CHr-CH, C 

HC ^C— CHa CH, HC C— COOH 

HC CH N gives on oxidation HC CH 

\/ I . \/ 

N CH, N 

Nicotine Nicotinic acid 



This formula for nicotine proposed by Pinner was confirmed several years later 
by Am6 Pictet's synthesis of this alkaloid. 

Physiological Action. — Nicotine is one of the most powerful poisons and scarcely 
inferior to hydrocyanic add in toxidty and rapidity of action. It appears to be 
toxic to all classes of animals. It is absorbed from the tongue, the eye and the 
rectiun even in a few seconds and from the stomach somewhat more slowly. 
Absorption of nicotine is also possible from the outer skin. Elimination takes 
place through the lungs and kidneys. In concentrated form nicotine is a local 
irritant, though, owing to the rapidity of its toxic action, it does not behave like a 
true corrosive nor does it cause inflammation of the mucous lining of the stomach 
even after a lethal dose. Nicotine, after causing stimulation for a brief period, 
then paralyzes the central nervous system and spinal cord, finally affecting vari- 
ous organs such as the heart, eyes and intestinal tract. Its poisonous influence 
probably extends to all parts of the brain, medulla oblongata and spinal cord. 
Huchard states that nicotine causes a general convulsion of the circulatory sys- 
tem which is apparent in chronic nicotine poisoning. In chronic tobacco poison- 
ing the general condition of health is disturbed and quite frequently the eyes are 
affected. In acute nicotine poisoning death ensues from paralysis of the respira- 
tory center. An action upon the heart is also always in evidence even in non- 
fatal cases. 



ine, tertiary cyclic amines, as pyridine and quinoline, also give similar iodo- 
methylates which are ammonium iodides with quinquivalent nitrogen: 

H H 

C C 

CH,v III ™r\V /\ /\ 

CHj^N +CH,.I = pS'UN.I; HC CH HC CH 

CH,/ ^g;/ I II +CH,.I= I Jj 

Trimethylamine HC CH HC CH 

Y V 

Pyridine / \ 

H,C I 



88 DETECTION OF POISONS 

Detection ol Wicodne 

Ether or low-boiling petroleum ether will extract nicotine 
from an aqueous alkaline solution. Spontaneous eviqx>ration 
of the solvent leaves the alkaloid as an oQy liquid having the 
odor of tobacco and a strong alkaline reaction. General alka- 
loidal reagents will precipitate nicotine from quite dilute scdu- 
tions, in which respect this alkaloid is very different from coni- 
ine« Phospho-molybdic add and potassium bismuthous iodide 
precipitate nicotine even in a dilution of i : 40,000; potas- 
siimi mercuric iodide in 1:15,000; gold chloride in 1:10,000; 
and platinum chloride in i : 5000. 

I. Cxystallizatian Test — Evaporate nicotine on a watch 
glass with a few drops of concentrated hydrochloric add. This 
will \ield a yellow, varnish-like residue which microscoi^c ex- 
amination will show to be entirdy amorphous (distinction 
between nicotine and coniine). If kept for a long time in a 
desiccator over sulphuric add, it will become indistinctly 
cr>*stalline. 

a. Roossin^s Test — Dissolve a trace of nicotine in ether, 
using a dr>* test-tube* Add to this solution about the same 
volume of ether containing iodine. Stopper and set the test- 
tube aside. The mixture will become turbid and deposit a 
browni;Sih nxl resin which will gradually become crystalline. 
After some time, ruby-red needles with a dark blue reflex will 
crvstalUze fr\^m the ether. These are •' Roussin's crvstals." 
If nianine is old or resinous* it will not as a rule give these 
crystals. 

3* Melxer^ Test*^ \i ^ dr\^p oi niv>>tine is heated to boiling 
with •* ^< o\\ of qxiv hlor\^h\ drin.* the mixtiire becomes dis- 
tinctly rx\l ri\is lot a\^|xlie^l to vvuiine causes no color. 

K>vlri«. V U.vOU^ V Uri v U,v U U a ^^^kMVc^ tk^uxt i»^^S* ia ntcr and 



NON-VOLATILE POISONS 



89 



I 



4. Schindelmeiser's Test.' — If nicotine that is not resinous 
is treated first with a drop of fonnaldehyde solution free from 
formic acid and then with a drop of concentrated sulphuric acid, 
the mixture takes on an intense rose-red color. If nicotine and 
fonnaldehyde are in contact for several hours, the solid residue 
obtained gives even a finer color reaction with a drop of nitric 
add. Only a little formaldehyde should be used, otherwise 
the solution becomes green after a while and decomposition 
takes place. 

Under the same conditions trimethytaniine, piperidine, pyridine, picoline, 
quinoline and aniline gave no color. Nor did extracts from putrefying horse-flesb 
and the entrails of animals, poisoned by arsenic or mercury, give the test, at least 

t when these extracts were prepared according lo the Stas-Otto method- 

5. Physiological Test. — When very small quantities of nico- 
tine are present, the physiological test should accompany the 
chemical tests. A very characteristic picture is given by frogs 
after administration of small doses of nicotine. First there is 
stimulation, then paralysis of the brain and respiratory muscles 

id apparent curare-action (tetanic con\-ulsions) . Thetoxic 
:tion of pure nicotine should be studied first. The experiment 
th a frog's heart, which shows temporary cessation of diastole, 
also very characteristic. 

AMLINE 

) Aniline, C»Hj.NH], upon evaporation of the ether extract from 

iie alkaline solution, will usually appear as reddish or brownish 

globules. Dissolve some of this residue in water and apply 

the aniline tests already described on page 45. A further 

I test for aniline consists in mixing some of the residue with a few 
^ops of concentrated stilphuric acid, and adding a few drops 
of potassium dichromate solution. If aniline is present, an 
evanescent blue color will appear. 



VERATRINE 



Pure offidnal v 
tlif composition CjiH„NOi. These a 



mixture of two isomeric alkaloids having 
e cevadine, also called crystallized vera- 



> Pharmueutische Zentral-Halle 40, 703 (1SQ9). 



90 DETECTION OF POISONS 

tritte* which is nearly insoluble in water; and amorphous veratridine which is 
^uhle in water. Even small quantities of the crystalline alkaloid will render 
v«catridine insoluble in water. On the other hand veratridine will prevent 
CYvadine from crystallizing. Consequently the crystalline base cannot be iso- 
lAied by recrystallizing officinal veratrine from alcohol or from any other solvent; 
iK>r can the water-soluble alkaloid be obtained by simple extraction with water. 

Sitftrttioii of Cevadine and Veratridine.— £. Schmidt uses the following 
m^hixi to isolate the crystalline and the water-soluble veratrine from officinal 
vtCAtHne. Place the officinal preparation in a beaker and dissolve in strong 
aK\>)h>1< Heat this solution to 60-70° and add enough warm water to produce a 
kwrmanent turbidity. Cautiously add just enough alcohol to clear the solution 
aihI aU^>w evaporation to take place slowly at 60-70°. A white, crystalline pre- 
^^^Ult» will presently appear. Filter with suction, wash the precipitate with a 
\^%%\^ \\\\\\tt alcohol and recrystallize from hot alcohol. This is crystalline vera- 
li^^'v i'lcNir the filtrate from the crystalline precipitate by adding a little alcohol 
Ai^t <^vA|H\rate at 60-70°. This will give a second crop of crystals. By repeating 
tKii( i\r\Ht'iui several times one may obtain in a crystalline condition about one- 
iVrvt «^ (^^ veratrine taken. Finally evaporate the filtrate from the crystalline 
x^>«siUl at the given temperature until there is no longer any odor of alcohol. A 
t\'^Bi(\lKAUIe quantity of a resinous mass which is a mixture of both alkaloids will 
^vAtAl^ The aqueous filtrate from this deposit will contain veratridine which 
Y^M tst \^M«lneii by rapidly evaporating the solution in vacuo over sulphuric add. 

l^yM^ttiM of Officinal Veratrine.— Veratrine appears as a \fhite, amorphous 

yy^t^>^ x^hloh is crystalline under the microscope. It has a sharp, burning taste 

MtA iKf minutest quantity introduced into the nostrils excites protracted sneezing. 

\i » Alm\wt insoluble in boiling water and the aqueous extract always has a 

^i^^^^N 4^\ltaU«c reaction; fairly soluble in ether (i : 10), benzene, petroleum ether 

HhA 4kiA\\ «UH>hol; and freely soluble in alcohol (i 14) and chloroform (i :a). 

Vi^ itW^ ik«\hitlon9 have a strong alkaline reaction. Officinal veratrine melts at 

s v> Sk^"^ I^^ * yellowish liquid which solidifies to a transparent, resinous mass. 

V tV Vv^^tfinc solution is faintly acid, ether will extract a very little of the alka- 

■V*»A \^V^^^ ^^^^ same conditions, chloroform and amyl alcohol will extract 

■u»>ry» ^1h* 4^lkaloid is usually deposited from ether as a white, amorphous pow- 

»V»- V*^\s«^\ho*molybdic acid, iodo-potassium iodide, tannic acid and potassium 

■***«* s¥^ i\^Md<> give distinct precipitates with an aqueous veratrine solution 

ss*-^»M»?Wi K>'^lr\>chloric add and diluted i : 5000. Chlorides of gold and plati- 

■i.«.u ^V>^ |sK He acid fail to show the alkaloid in this dilution. 

VVUMffrtH^' " * ^****^'^^ ^^ saturated barium hydroxide, or alcoholic |>otas- 
^ViJu Vs^N\\<\l<» solution, crystallized veratrine (cevadine) is hydrolyzed into 
» i^Uiv ««s««^ «^»>^^ Irvine: 

C^fHiiNOi + H2O = CfcHsOa^ + C27H4,NO» 
CtVAdinc Angelic Cevine 

acid 

V'W*^*S 4k\i\\ vO «^nd tiglic add (II) are stereo-isomers: 



I iMIi— C— H II. H— C— CH,^ 

-COOH CH,— C— COOH 



\ 111 ^ •" 



NON-VOLATILE POISONS 91 

M. Freund^ has shown that cevadine takes up only one acetyl or benzoyl 
group, whereas cevine takes up two. The following formulae show these 
relationships: 

/0.C^H70 yOH 

X)H N)H 

Cevadine Cevine 

i 1 

/O.CiHyO /O.CO.CH, 

C,tH4iN0< C,7H4iNO.< 

X).CO.CH, X).CO.CH, 

Acetyl-cevadine Diacetyl-cevine 

By means of hydrogen peroxide M. Freund has converted cevine into cevine 
oxide, Cs7H4sNOb, which crystallizes well and contains one more atom of oxygen. 

y 

This compound must belong to the class of the amino-oxides, i^vN >■ O, for sul- 
phurous add easily converts it into cevine. 

Detection of Veratrine 

1. Concentrated Sulphuric Add Test — Poiir a few drops of 
concentrated sulphuric acid upon a trace of veratrine. The 
alkaloid will have an intense yellow color and, if stirred, will 
give a solution of the same color. Gradually this color will 
change to orange, then to blood red and finally to cherry red. 
(Jentle heating will hasten this color change and veratrine, dis- 
solved in concentrated sulphuric acid, will give a fine chferry red 
solution almost immediately. 

Frohde's and Erdmann's reagents give color changes similar 
to those caused by sulphuric acid. 

2. Concentrated Hydrochloric Add Test— If a trace of vera- 
trine is dissolved in i or 2 cc. of cold concentrated hydrochloric 
acid, the solution will be colorless. When this solution is 
heated 10 to 15 minutes in a boiling water-bath, a cherry red 
color will appear. This color will last for a day and even 0.2 
mg. of veratrine will produce it. 

3. Concentrated Nitric Acid Test — Concentrated nitric acid 
dissolves veratrine with a yellow color. 

4* Weppen's Test — Thoroughly mix in a mortar i part of 
veratrine with about 5 parts of finely powdered cane sugar. 
Add a few drops of concentrated sulphuric acid to some of 

^Berichte der Deutschen chemischen Gesellschaft 37, 1946 (1904). 



92 DETECTION OF POISONS 

the mixture upon a watch glass. At first a yellow color will 
appear and later, beginning at the margin this will change to 
grass green and finally to blue. Breathing upon the mixture 
will cause the color to change more quickly. Too great an 
excess of cane sugar must be avoided. 

E. Laves^ substitutes an aqueous furfurol solution for cane 
sugar in this test. Mix in a test-tube 3 or 4 drops of i per cent. 
aqueous furfurol solution with i cc. of concentrated sulphuric 
acid; Add 3 to 5 drops of this solution to the substance to be 
tested so that it just touches the edge of the liquid. If veratrine 
is present, a dark streak will gradually run from the substance 
into the liquid. At the starting point it will appear blue or blue 
violet and farther away green. If substance and liquid are 
stirred with a glass rod, the liquid will become dark green. 
After some time, or more quickly when warmed, the color 
will become blue and finally violet. 

5. Grandeau's Test. — Direct addition of 1-2 drops of bro- 
mine water to the yellow solution of veratrine in concentrated 
sulphuric acid produces an immediate purple color almost iden- 
tical with that appearing when the solution of the alkaloid in 
concentrated sulphuric acid stands a long time or is gently 
warmed. 

6. Vitali's Test — Dissolve veratrine in a few drops of fuming 
nitric acid and evaporate the solution to dryness upon the 
water-bath in a porcelain dish. A yellowish residue will remain. 
If this is cooled and then moistened with an alcoholic potas- 
sium hydroxide solution, the color will change to orange red or 
red violet and stirring will produce a solution having the same 
color. 

Atropine, hyoscyamine, scopolamine, as well as strychnine, 
respond to this test in a very similar manner. 

STRYCHNINE 

Strychnine, C2iH2aNa02, occurs with brucine chiefly in nux vomica and 
Ignatius beans, constituting the larger part of the mixed alkaloids. The former 
contains 2.93-3.14 per cent, of these two alkaloids and the latter 3.1 1-3.22 per 

1 Pharmaceutische Zeitung, 37, 338. 



NON-VOLATILE POISONS 



93 



The free base strychnin 
thomfaic system which mell a 
cold and 3500 parts of hot wal 
taste. It is □early insoluble 11 
solves in t6o parts of cold and i 



; forms colorless, shining prisms belonging to the 
268°. The alkaloid dissolves in 6600 parts of 

:r, giving alkaline solutions having a very bitter 
absolute alcohol and in absolute ether. It dis- 

! parts of boiling alcohol (50 per cent, by volume) ; 
it readily in chloro- 



it is also soluble in commercial ether and in benzene; but 
form (6 parts at 13°). Strychnine diluted with water i ;6oo,c 
by its bitter taste. 

Strychnine is a monacid base combining with one equivalent of add and fona- 
iog salts which are usually crystalline. These salts have a very bitter taste and 
are very poisonous. The best known strychnine salt, and one used medicinally, 
u the nitrate, C,iH,iN,Oj,HNOi. The combination of one molecule of strych- 
nine with one molecule of an alkyl haloid, for example, methyl iodide, to form 
■trychnine iodo-me thy late, CiiRijNO,.CHi.I, shows that the alkaloid is a tertiary 
base. Sodium methylate (CHi.ONa) in alcoholic solution converts strychnine 
into strychnic acid which is probably an imino-carboiylic acid. Strychnic add 
loses a molecule of water, when its solutions are boiled in presence of mineral 
adds, and is changed to strychnine. Because of this behavior Tafcl regards 

Fihoine as an inner anhydride of strychnic add, one containing a group oi the 
sctec of an add imide: 
a 



4 



fOn the basis of Tafel's strychni 
npressed as follows: 



} + H,0 i± (C„H„0)J-CO0H 
Stt]ichnic acid 

formula, strychnine iodo-metbylate would be 



(C,oH„0)- 



N<; 



Pbysiologics] Action. — Strychnine increases reficx irritability of the brain and 
(pinal cord. Even ihe slightest stimulus, espedally if acoustic, optical, or tactile, 
may cause powerful reflexes after large doses of this alkaloid. Convulsions 
may followeachatimulus.it the dose is sufiident. Very large doses of si rychnine 
cause curare-like paralysis of the peripheral ends of motor nerves in frogs and 
other warm-blooded animals. It may also affect the musdes of the heart. 
Strychnine diminishes the motile power of leucocytes and then arrests their 
motion. The poison also affects plant protoplasm, at least that of Mimosa 
pudica, in that the plant's motor organs lose their elasticity and fiexibility. 

Aside from the saliva, bile and milk, the urine is the main channel Ibrough 
which strychnine is eliminated from the organism. Human urine may contain 
even the unaltered alkaloid. ElirnJnation begins during the first hour, is slight 
after 1 days but is not complete until much later. Mote unaltered strychnine is 
eUminated after large than alter small doses. In the former case 70-7S pM «nt. 
of the alkaloid may remain undecomposcd. The liver, kidneys, brain and spinal 
cord may store up unchanged strychnine. (See R. Robert, "lnto»tkationen.'*) 



94 DETECTION OF POISONS 

Detection of Strychiiiiie 

Sodium and potassium hydroxide, anmipnia and alkaline 
carbonates precipitate the free base strychnine from aqueous 
solutions of its salts as a white crystalline solid: 

C,iH„N,0,.HNO, + NaOH = C,iH„N,0, + H,0 + NaNO,. 

Ether will extract strychnine from an alkaline solution and 
deposit the alkaloid on evaporation in fine crystalline needles. 
Chloroform takes up the alkaloid more freely, since strychnine 
is considerably more soluble in this solvent than in ether. Even 
very dilute solutions of strychnine salts give precipitates with 
most of the alkaloidal reagents. Tannic add, potassium mer- 
curic iodide and phospho-tungstic acid produce white precipi- 
tates; gold chloride and phospho-molybdic acid yellow precipi- 
tates; and iodo-potassium iodide brown precipitates. To 
obtain tests with these reagents, the residue from ether should 
first be dissolved in very dilute hydrochloric acid. 

Concentrated sulphuric acid, Erdmann's and Frohde's 
reagents dissolve perfectly pure, brucine-f ree strychnine without 
color. 

Strychnine is soluble in concentrated nitric acid with a yellow- 
ish color. Potassium dichromate, added to solutions of strych- 
nine salts, precipitates strychnine dichromate, (C2iH22N202)2.- 
H2Cr207, in the form of fine yellow crystalline needles which 
upon recrystallization from hot water appear as shining orange- 
yellow needles. 

Potassium ferricyamde, added to solutions of strychnine 
salts, precipitates golden-yellow, crystalline strychnine ferri- 
cyanide (C2iH22N202)2.H8Fe(CN)6 + 6H2O. 

Special Reactions 

I. Sulphuric Acid-Dichromate Test — Dissolve a very little 
strychnine in 2 or 3 drops of concentrated sulphuric acid 
upon a watch glass. The solution should be colorless. Add a 
fragment of potassium dichromate and hold it firmly in one place 
upon the glass. Intense blue or blue-violet streaks will come 
from the potassium dichromate, if the watch glass is tilted up 



NON-VOLAnLE POISONS 



95 



and down. If the entire mixture is stirred, the sulphuric acid 
will have a beautiful evanescent blue or blue-violet color. 

This test may also be made by scattering upon the surface of 
the solution of strychnine in concentrated sulphuric acid a few 
particles of coarsely powdered potassium dichromate and mix- 
ing well with a glass rod. In this way the blue to blue-violet 
color reaction is given very beautifully. The blue color is not 
permanent. It soon changes to red and finally to dirty green.' 

Strychnine chromate and ferricyanide give this test especially 
well. To prepare the former salt, pour a very dilute potassium 
dichromate solution over strychnine upon a watch glass. When 
the two substances have been in contact for some time, pour 
the remaining liquid from the strychnine chromate. Wash 
the precipitate once with a little water. Put some strychnine 
chromate upon the end of a glass rod and draw it through a 
few drops of concentrated sulphuric acid upon a watch glass. 
This will produce violet and blue streaks in the acid. 

Mandelin's reagent,^ that is to say, vanadic-sulphuric acid, 
jJves this strychnine test very well. The blue or violet color 
pven by this reagent with strychnine is more permanent than 
3iat produced by potassium dichromate. The color finally 
lianges to orange-red. 

Other oxidizing agents may be substituted (or potassium dichromate, u 
potassium permanganate, lead peroxide, manganese dioxide, potassium ferri- 
cyanide (see above), cenum oxide and vanadic add (Mandelin's reagent). But 
ndther potassium nitrate nor nitric Bcid can be used, as these reagents even pre- 
vent this test. Consequently strychnine nitrate does not give the test. 

2. Physiological Test. — Dissolve the ether residue in a little 
very dilute hydrochloric acid. Evaporate the filtered solution 
to drjTiess upon the water-bath. Dissolve the residue in pure 
water (about i cc.) and inject this solution into the lymph sac 
on the back of a Uvely frog. Keep the experimental frog in a 
large, loosely covered beaker. Toxic symptoms will appear in 
S to 30 minutes, depending upon the quantity of strychnine. 

' According to Tafel (Annaleo der Chemie und Pharmaiie, 168, 133 (iSoO.thi* 
color reaction is characteristic of many anilides and is due to the presence o( the 
group -CO-N = . 

' See page J14 for the preparalioti of this reagent. 



96 DETECTION OF POISONS 

Strychnine does not increase reflex irritability for all kinds of 
stimuli but only for tactile, optical and especially for acoustic 
stiiriuli. When the dose of strychnine is sufficiently large, 
each kind of stimulus mentioned will produce convulsions like 
those caused by tetanus. For example, if the beaker contain- 
ing the "strychnine frog" is gently tapped, this slight acoustic 
stimulus is sufficient to produce convidsions. 

Detection of Strychnine in Presence of Brucme 

More than traces of brucine prevent detection of strychnine 
with concentrated sulphuric acid and potassium dichromate. 
Under certain conditions Mandelin's reagent will show strych- 
nine more or less distinctly in presence of brucine. Dissolve 
the ether residue in concentrated sulphuric add, if brucine is 
present, and add a trace of concentrated nitric add. A red 
color indicates brudne. When the color has changed to yellow, 
add a fragment of potassium dichromate and stir. The mixture 
will become blue or reddish violet, if strychnine is present. 

Solid potassium permanganate, stirred with concentrated sulphuric add alone, 
will give a dark green solution which is yellowish green in a thin layer and assumes 
with time a red to violet color on the margin. 

The same procedure used to estimate these two alkaloids 
quantitatively will permit detection of strychnine even in pres- 
ence of considerable brucine. Dissolve the residue containing 
brudne in about 2 cc. of dilute sulphuric acid, add 2 drops of 
concentrated nitric acid and let the mixture stand 4 hours. 
Render alkaline with excess of sodium hydroxide solution and 
extract thoroughly with ether. The residue from ether will be 
brucine-free or nearly so. Strychnine thus treated will give 
very satisfactory tests with concentrated sulphuric acid and 
potassium dichromate and with Mandelin's reagent. 

BRUCINE 

Brucine, C2sH2eN204, crystallizes in transparent, monoclinic prisms or shining 
leaflets. Crystals from water contain either 4 or 2 molecules and from alcohol 
2 moleciUes of water of hydration. It melts in its water of hydration only a few 
degrees above 100°, whereas the anhydrous base melts at 178^. Brucine b 
more readily soluble than strychnine both in water and in alcohol and therefore 



NON-VOLATILE POISONS 97 

15 dissolved in the motlier liquors from the preparation of strychnine. It 
is also more soluble than strychnine in ether. Brudne solutions have a very 
bitter taste and a strong alkaUne reaction. Benzene, but especially chloroform 
and amy! alcohol, are encellent solvents forhrucine. Brudne differs from strych- 
nine in bdng deposited usually amorphous by evaporation of its ether solution. 

Brudne is a monadd, tertiary base and as such forms addition products with 
one molecule of an alkyl iodide. For example, with methyl iodide it gives 
brucine iodo-mcthylate, Ci>H,,NO..N.CH,I. With one equivalent of add 
brudne gives in part crystalline salts. Brudne nitrate, CnHxNiOt.HNOi. 
»H,0, crystaUizes in rectangular prisms. 

Brucine may be shown by Zeisel's method' to contain two methoiyl groups 
(-OCH,). 

Heated in sealed tube to 8o° with sodium and alcohol, until solution is complete, 
bnidne is converted into brudc add, C)iH,iNiOi.HjO, which contains an imino- 
group C = NH) in its molecule since it forms a nilrosamioe, Tafel and Moufang' 
^^^ipress the relationship between brudne and brudc add as follows: 

^HSeate 

^BucicK 

■ Ethi 



C«H„(0CH,),O— CO + H,0 j=! CioH,„(OCH,),0— COOH 



LSeated with water, brudc add is converted into brudne. Consequently 
[c Jidd is related to brudne as stiychnic add is to strychnine. 



Detection of Brucine 

Ether, benzene or chloroform will extract brucine from an 
alkaline solution. Evaporation of the ether extract usually 
leaves the alkaloid in an amorphous condition. The sensitive- 
ness of the alkaloidal reagents toward brucine is as follows: 
lodo-potossium iodide (i : 50,000) Potassium bismuthous iodide (1 : jooo) 

Potassium mercuric iodide (i ijo.ooo} Phospho-molybdJc add (t : looo) 
Gold chloride {1 : 30,000) Tannic add (i :aooo) 

Platiuic chloride (i ; 1000). 

' Many alkaloids contain one or more, sometimes three or more, mcthoxy! 
groups {— OCHi) united nith a benzene nucleus. The determination of the 
number of such groups in the molecule is of the greatest importance as a step in 
ealabltshing the constitution of an alkaloid, because in this way some of the car- 
bon, oxygen and hydrogen atoms are at once disposed o{. The method employed 
for this purpose depends on the (act that all substances containing methoiyl 
groups are decomposed by hydriodic add, yidding methyl iodide and a hydroxy] 
compound. By estimating tbe methyl iodide obtained from a given quantity 
of a, compound of known molecular weight, it is easy therefore to determine the 
number of methoxyl groups in the molecule. This method was first applied by 
Zeisel and b of general application. (Perkin and Kipping, "Organic Chem- 
istry," page 498.) 

'Annaien der Chemie und Pharmazie 304, )S (iSqq). 



98 DETECTION OF POISONS 

1. Nitric Acid-Staxinous Chloride Test — Concentrated nitric 
acid dissolves brucine aad its salts with a blood red color. This 
color, however, is slightly stable and soon changes to yellowish 
red and finally, especially with heat, to yellow. . Add a few 
drops of freshly prepared, dilute stannous chloride solution 
to this yellowish red or yellow solution. An intense violet 
color will appear. Heat usually changes this violet color again 
to yellowish red, but addition of a few more drops of stannous 
chloride solution will cause the violet color to reappear. The 
smaller the quantity of nitric acid, the more likelihood that this 
test will give a good result. Colorless ammonium sulphide 
solution may be substituted for stannous chloride. 

2. R. Mauch's Modification of Nitric Acid-Stannotis Chloride 
Test. — An excellent result can be obtained with this test in the 
following manner. Dissolve brucine in 60 per cent, aqueous 
chloral hydrate solution and put about 0.5 cc. of this solution 
into a test-tube. Add very little dilute nitric acid and thor- 
oughly mix the two solutions. Add this mixture to 3 times 
its volume of concentrated sulphuric acid so that the former is 
on the surface. A yellowish red to deep red zone, depending 
upon the quantity of brucine, will appear immediately. When 
the upper layer becomes yellow, introduce by a pipette a little 
stannous chloride solution^ as a top layer. A brilliant, intensely 
violet zone will appear between the two upper layers. The 
intensity of this color will gradually increase, especially if the 
test-tube is gently tilted to and fro. 

ATROPINE 

H2C— CH CH2 CH2OH 

I I I 

N.CH, CH— O.CO— CH 

I I I 

H2C— CH CH2 CHfi 

Atropine, CnHasNOi, crystallizes in shining pointed needles which melt at 115* 
and dissolve in 600 parts of water, 50 parts of ether and 3.5 parts of chloroform. 
It is also soluble in alcohol, amyl alcohol and benzene. The aqueous solution of 

* Prepare stannous chloride solution by dissolving i part of stannous chloride 
in 9 parts of hydrochloric acid having a specific gravity of 1.12 (about 24 per 
cent. HCl). 



NON-VOLATILE POISONS 99 

the alkaloid is alkaline and has a lasting, unpleasant, bitter taste. Unlike the 
optically active hyoscyamine, atropine is inactive. 

Constitutioii. — Heated with hydrochloric acid at 120-130**, atropine is decom- 
posed into tropic acid and tropine: 



H 



H 
H,C C CH, 



I 
H,C— N HC— O 



H,C C CH, 

H 



i 



OH 



— C— CH 
CeH. 



H 
CHx— OH H,C C CH, 



H,C— N HC— OH + 



H,C C CH, 

H 

Atropine Tropine 

CHi— OH 

I 
HO—C— CH 

'I i 
O C«H» 

Tropic acid 

Heated with barium hydroxide solution, atropine yields atropic acid which is 
unsaturated and differs from tropic acid by one molecule of water: 

CH,.OH CH, 



I 

CH.C«H» - H,0 



A 



I 

C.CeHt 



OOH COOH 

Tropic acid Atropic acid 

Since the structure both of tropine and tropic acid has been determined by 
synthesis as well as by decomposition, that of atropine is also known. Nitro- 
gen in atropine is in the tertiary condition. Hyoscy amine is the stereo-isomer of 
atropine. The former, heated at 110° out of contact with air, or allowed merely 
to stand in alcoholic solution with addition of a few drops of an alkaline hydroxide 
solution, is changed to inactive atropine. Atropine most likely is the racemic 
form, whereas hyoscy amine is the laevo-rotatory modification of this isomeric 
base. The degree of rotation of hyoscyamine is fa]D = —20.97**. Toward 
alkaloidal reagents and when heated with concentrated sulphuric acid, hy- 
oscyamine behaves like atropine. It also resembles the latter in giving Vitali's 
reaction (see below). 

Putrefactioxi. — Ipsen* has found atropine very resistant in presence of putre- 
fying material. Even after 2 years he could detect the alkaloid which had been 
exposed to the influences of decomposition. He experimented with 0.05 gram 
of atropine sulphate in respectively 300 cc. of blood, urine and beer and with pure 
atropine in 300 cc. of blood. 

^ Vierteljahrsschrift ftir gerichtliche Medizin und offentliches Sanity tswesen, 
3I1 308. 




100 DETECTION OF POISONS 

Detection of Atropine 

Ether, benzene or chloroform will extract atropine from a 
solution alkaline with sodiimi hydroxide or carbonate solution. 
In a special search for atropine use sodium carbonate solution 
and extract with chloroform which is a better solvent than ether. 
Evaporate the solvent and test the residue, which is usually 
amorphous, as follows: 

1. Vitali*s Test. — Dissolve the alkaloid in a few drops of 
fuming nitric acid, and evaporate the solution in a porcelain 
dish to dryness upon the water-bath. Moisten the yellowish 
residue when cold with a few drops of a solution of potassium 
hydroxide in absolute alcohol. An evanescent violet color 
will appear, if atropine is present. 

Hyoscyamine and scopolamine also give Vitali's test. Strychnine and vera- 
trine behave similarly. This test therefore is characteristic of the atropine 
alkaloids only in the absence of the two latter alkaloids. 

2. Odor Test — Heat a little atropine in a dry test-tube until 
a white vapor appears. An agreeable odor will arise at the same 
time. Then add about i cc. of concentrated sulphuric acid, and 
heat until the acid begins to darken. Dilute at once with 
about 2 cc. of water. During the foaming there will be an in- 
tense, sweetish odor like that of honey. By this test, which 
was formerly the only method of identifying atropine, o.oi 
gram of the alkaloid can be detected. 

3. Ph3rsiological Test. — Atropine acts in a very characteristic 
manner upon the pupil of the eye, and this behavior can be 
employed as a test. One drop of an atropine solution diluted 
1 : 130,000 will produce a noticeable enlargement of the pupil. 
Dissolve a small portion of the ether residue in 4 or 5 drops of 
very dilute sulphuric acid, and introduce a drop of this solution 
into a dog's or a cat's eye. The enlargement of the pupil often 
persists for several hours. The utmost care should be taken 
in performing this test, if applied to the human eye. 

The following alkaloidal reagents are especially sensitive 
toward atropine: iodo-potassium iodide, phospho-molybdic 
acid (1:10,000), gold chloride, phospho-tungstic acid, potas- 



NON-VOLATILE POISONS 



101 



saaa mercuric iodide, potassium bismuthous iodide. Picric 
acid, added to solutions of atropine salts that are not too 
dilute, will precipitate atropine picrate as yellow leaflets, 
[ Plattnic chloride gives monoclinic prisms. 



I 



HOMATROPINE 

Homstropinc, CiiHuNOj, is the Iropyi ester of phenyl-glycob'c or mandelic 
add. The hydrochloride of this base is obtained hy heating a mixture of tropine, 
W-Q—CJi CH C H mandelic acid and hydrochloric acid, the latter 

T I acting at a dehydrating agent. 

CH.CX 



N.CH, C 

I 



-in. 



fiiC— CH- 

u strong as that of the na 
whereas [hat of atropine 
Rtropine, Homatropine i: 
■cids. This alkaloid does 



The hydrobromide of homatropine (CuHti- 



OH 



NOi.HEr) b used in medidne as a substitute 
for atropine. Its action on the pupi! is nearly 
tural alkaloid and its eSect disappears In 13-14 hours, 
often lasts 8 days. Moreover it is less toiric than 
a strong tertiary base which forms neutral salts with 
Vilali's test. It melts at 92-56"; hyoscya- 



cocAnra 



mine at 108°; and atropine a 

Offidnal homatropine hydrobromide may be distinguished from the hydro- 
bromides of atropine and hyoseyamine by warming the substance in a test-tube 
with a little chloroform. This solvent dissolves the latter two salts in every pro- 
portion but homatropine hydrobromide is insoluble. An alternative procedure 
consists in dissolving the given salt in a little water, precipitating the base with 
■odium carbonate solution, and extracting with ether. Dehydrate the ether 
extract with potassium carbonate and evaporate slowly in a moderately warm 
place. This method will give crystals of the alkaloid. Dry these crystals in 
tacito over concentrated sulphuric add and determine their melting point. 

» Cocaine, CitHhNOi, crystallizes from alcohol in large, colorless, monoclinic 
lams which mell at 98°. It has a bitterish taste and, placed upon the tongue, 
C CH CH COOCH causes temporary, local aniesthesia. The 

I I I alkaloid is only slightly soluble in water 

I N.CH. CH— OOC— CH, (1:700), but easUy soluble in alcohol, ether, 
I j_ chloroform, benzene and acetic ether. Its 

' solutions are strongly alkaline and Itevo-rota- 

tory. Dilute adds easily dissolve cocaine and in most cases form readily erys- 
tallixable salts. The fixed alkalies, ammonia and alkaline carbonates predpitate 
the free base from solutions of its salts. 

Coaatitution.^ — Cocaine is a monocid, tertiary base, since it adds a molecule of 
CH|I. On distillation with barium hydroride. this alkaloid loses methyl amine 
(CHi.NHi), thus proWng the attachment of a methyl group to nitrogen. Co- 
caine must therefore contain the group = N— CHi, This base is also the 
methyl ester of an add and at the same lime the benzoyl derivative of an alcohol, 
B<ior it is decomposed into benzoyl- ccgonine nnd methyl alcohol when heated with 



,C— CH— 



102 



DETECTION OP POISONS 



water. If mineral adds, barium hydroxide or alkalies are used instead of water, 
the primary product, benzoyl-ecgonine, is further decomposed into ecgonine, 
benzoic acid and methyl alcohol. Taking the structural formula proposed by 
WillstS,tter, we may express this reaction as follows: 



Cocaine 
H,C CH, 



CH, 

I 
HC— N— CH 

I H 



Bcgonine 
H,C CH, 

I I I 
HC— N— CH 



Bensoic 
acid 

C,H, 



Methyl 
alcohol 

CH. 



CeH^CO 



HiC— C— CH 

-0 CO— bCH, 
HOH HO :H 



I " I 
= H,C— C— CH + 

I I 



CO 

i 

H 



A 

H 



+ 



O 

I 
H 



Water 



CO 

I 

o 

H 



Ecgonine (I) heated with phosphorus oxychloride loses a molecule of water and 
passes into anhydro-ecgonine (II). The latter heated to 280° with fuming 
hydrochloric acid loses carbon dioxide and is converted into tropidine (III). 
Tropidine heated with a caustic alkaline solution adds a molecule of water and 
passes into tropine (IV), the basic cleavage product of atropine. Evaporation of 
tropine with tropic acid in dilute hydrochloric acid solution yields atropine (V). 
Thus it is possible to start with the alkaloid cocaine and synthesize the alkaloid 
atropine. The series of changes involved is as follows : 
H,C— CH CH.COOH H,C— CH CH. jCOO: H 



, CH. 



N.CH 



H,C— CH CH 

Bcgonine (1) 

H,C— CH CH, 



OH 
H 



-H,0 

► 



[, C] 



N.CH, CH 

ii 

H,C— CH CH 



-CO, 



N.CH, CH 

I I 

H,C— CH CH 

Tropidine (III) 

H,C- 



+ H,0 



Anhydro-ecgonine (II) 

H,C— CH CH, 

I 



CH,.OH 



-i 



N.CH, CH.O: H + HO OC-CH 

I 



CH- 



H,C— CH CH, 

Tropine (IV) 

-CH, CH,.OH 



CeHf 
Tropic acid 



H,C— CH 



N.CH, CH.O.CO.CH 
-CH, 



+ H,0. 



CeHi 



Atropine (V) 

Behavior in the Animal Organism. — Experiments upon dogs and rabbits show 
that the former animal eliminates through the kidneys not more than 5 per cent, 
of the cocaine as such and the latter none at all. As the urine of these animals 
also contains no ecgonine, the supposition is that the alkaloid is profoundly 
changed in the animal organism. The same is true of the human organism. 
Proells^ was able to detect cocaine in cadaveric material at most after 14 days. 
In the living organism the alkaloid is said to be changed rapidly into ecgonine. 

* Apotheker-Zeitung 16, 779, 788. 



NON-VOLATILE POISONS 103 

Detection of Cocaine 

Ether, chloroform or benzene will extract cocaine from an 
alkaline aqueous solution. Most of the alkaloidal reagents will 
precipitate cocaine even from very dilute solutions of its salts. 
The reagents especially sensitive are: iodo-potassium iodide, 
phospho-molybdic and phospho-tungstic acids, potassium mer- 
curic iodide, potassium bismuthous iodide, gold and platinum 
chlorides, and picric acid. 

Pure concentrated sulphuric and nitric acids, as well as Erd- 
mann's, Froehde's and Mandelin's reagents, dissolve cocai^ie 
without color. 

1. Precipitation Test. — If i or 2 drops of potassium hydroxide 
solution are added to an aqueous solution of a cocaine salt not 
too dilute, it will become milky. First, resinous globules and 
later fine, crystalline needles of the free base, cocaine (melting 
point 98**), separate from solution: 

Ci7H,iN04.HCli + KOH = C17H21NO4 + H,0 + KCL 

In applying this test to the ether residue, dissolve a consider- 
able quantity in a few drops of dilute hydrochloric acid and add 
potassium hydroxide solution drop by drop until alkaline and 
cool well by setting in ice. Special care must be taken to have 
the alkaloid pure enough when dry for a melting-point deter- 
mination. 

This test is not characteristic of cocaine (except the melting 
point which, however, requires considerable pure material), 
because most of the alkaloids are precipitated by potassium 
hydroxide solution in much the same way. 

2. Potassium Permanganate Test — Add saturated potassium 
permanganate solution drop by drop to a concentrated aqueous 
solution of a cocaine salt. This reagent will give a violet, 
crystalline precipitate of cocaine permanganate. In applying 
this test to the ether residue, dissolve a considerable quantity 
in 2 drops of dilute hydrochloric acid and evaporate the solution 

* Cocaine hydrochloride crystallizes from a concentrated aqueous solution in 
fine prisms containing 2 molecules of water which are easily given off. This salt 
crystallized from alcohol is anhydrous and has the formula Ci7HaiN04.HCl. 
The anhydrous compound is the officinal salt. 



104 DETECTION OF POISONS 

upon the water-bath. Dissolve the residue in as little water 
as possible and add potassium permanganate solution. 

3. Chromic Acid Test — ^Add a few drops of a 5 per cent 
chromic acid solution, or potassium dichromate solution of 
corresponding concentration (7.5 per cent) to a solution of a 
cocaine salt. Each drop will produce a precipitate which will 
immediately disappear if the solution is shaken. Then add 
to the clear solution about i cc. of concentrated hydrochloric 
acid which will produce an orange-yellow precipitate more or 
less crystalline. 

4. Detection of Benzoyl Group. — This test requires at least 
0.2 gram of cocaine. First, digest the cocaine a few minutes 
in a test-tube with 2 cc. of concentrated sulphuric acid upon a 
boiling water 'bath. Cool and dilute with a little water, all 
the while keeping the mixture cold. A white cr3rstalline pre- 
cipitate of benzoic add will appear. Collect and dry this 
precipitate upon a filter. Benzoic acid may be recognized by 
subliming the precipitate, or, if the quantity is 'sufficient, by 
determining the melting point (120°). 

Benzoic acid may also be extracted with ether. Mix the 
residue, obtained by evaporating the solvent, with i cc. of 
absolute alcohol and the same quantity of concentrated sul- 
phuric acid. The characteristic odor of ethyl benzoate, 
CeH5.CO.OC2H6, will be recognized. 

5. Reichard's^ Test — Addition of a concentrated aqueous 
solution of sodium nitroprusside, Na2Fe(CN)6N0.2H20, drop 
by drop to a cocaine salt solution, containing at least 4 mg. 
of cocaine per cc, causes an immediate turbidity which will 
appear under the microscope as well formed reddish crys- 
tals. These crystals will dissolve, if the liquid is warmed, and 
appear again if the solution is well cooled. Morphine does not 
give this test. 

6. Goeldner's^ Test — Mix about 0.005 gram of pure resor- 

^ C. Reichard, Chexniker-Zeitung 28, 299 (1904). Pharmazeutische Zeitung 
Z904, Nr. 29. Pharmazeutische Zentralhalle 45, 645 (1904). 

' Pharmazeutische Zeitschrift ftir Russland 28, 489 and Zeitschrif t fQr analy- 
tische Chemie 40, 820 (1901). 



NON-VOLATILE POISONS 105 

inol (CgH4(0H)i I, 3) in a small porcelain dish with 5-6 drops 
i pure concentrated sulphuric acid. Add about o.oz gram of 
e hydrochloride to this solution which usually has a faint 
Yellowish color. There is a vigorous reaction, during which the 
liquid acquires a fine blue color like that of the corn flower. 
' The intensity of tliis color gradually increases. Sodium hydrox- 
ide solution will change the blue to Hght pink. 

7. Physiological Test. — Dissolve the material (the residue 

from the ether extract) in a few drops of dilute hydrochloric 

acid and evaporate the solution to dryness upon the water-bath. 

Dissolve the residue in a little pure water and apply this solu- 

I tion to the tongue. Cocaine produces a temporary anesthesia. 

R. Kobert (" Intoxikationen ") has found small frogs suffi- 

I dently sensitive for use in the physiological test for cocaine. 

rXhe effects to be observed are dilatation and fixedness of the 

[ pupil, enlargement of the palpebral fissure and also stimulation 

I of the nervous system. Administer the same quantity of co- 

I caine hydrochloride to animals for comparison. 



I 



PHYSOSTIGMINE 

Physostigmine, CnHiiNiOi, also called eserine, occurs in the Calabar bean, 
the seed of Physoatigma venenosum. This alkaloid is deposiied from benxene 
solution upon spontaneous evaporation oF the solvent in large, apparently 
tbombic crystals melting at 105°. Though but slightly soluble in water, it 
dissolves Ereely in alcohol, ether, benzene or chloroforin. Pbysostigmine solu- 
doDs are strongly alkaline, almost tasteless and tsvo-rotatory. It is a strong 
mooadd tertiary base, forming salts with adds that easily undergo decomposition 
uid crystallize with difficulty. Light and heat cause acid and alkaline solutions 
of this alkaloid to turn red. Owing to this tendency of physosUgmine to undergo 
decomposition, care must tie taken during its isolation to keep it from light and oic 
Uid also to avoid rise of temperature. Exclusion as far as possible of free min- 
eral adds and caustic alkalies is a bo desirable. 



Detection of PhysoBtigmine 

Concentrated sulphuric and nitric adds dissolve pbysostigmine with a yellow 
color which soon changes to olive-green. The alkaloid evaporated upon the 
water-batb with fuming nitric acid leaves a residue having a green margin. 
Water, alcohol and sulphuric add dissolve this residue with a green color. 

1. Ammoiiia Test — If a small quantity of a pbysostigmine salt is evaporated 
to dryness upon the water-bath with Brnmonium hydroride solution, a blue or 
t^lue-green residue will remain. This will dissolve in alcohol with a blue color. 



106 DETECTION OF POISONS 

Excess of dilute mineral mdd. or acetic acid, added to this sdutioii will diange the 
color to red. The solution is also strongly fluorescent. Fiamined spcCtio- 
scopically the blue alkaline solution shows one abfioq>tioii buid in the red; and 
the red acid solution one absorption band in the yellow. 

A drop of concentrated sulphuric add, added to the Uue residue from evapora- 
tion with ammonia, will g;ive a green solution. The green odor diluted with 
alcohol wUl change to red. If the alcohol is evaporated, the green color wiD 
reappear. 

2. Rub res e rin e Test — If an aqueous solution of a physostigmiDe salt is shaken 
for some time with an excess of potassium or sodium hydroxide sdutioo, a red 
coloring matter, rubreserine (CisHi«XsO«), is formed. This cotnpound separates 
as red needles which become greenish blue on further oxidation owing to formation 
of eserine blue. 

Barium hydroxide solution may be substituted for the caustic alkali. This 
reagent first produces a white precipitate which soon becomes red on being shaken. 
Sometimes this change occurs even in the cold but invariably takes place with 
heat. 

8. Physiological Test — The marked action of ph>'sostigmine in causing con- 
traction of the pupil is ver>' characteristic. It b advisable to use the cat's eye 
for this test. Even o.i mg. of this alkaloid wiU produce noticeable contraction. 

CODEINE 

Codeine, Ci7Hi8^CHs)XOs, the methyl ether of morphine, crystallizes from 
water, or from ether containing water, in colorless, tran^>arent octahedrons 

CH which are often very large. These cr>'stab are quite 

H Hs I easily soluble in water. One part of the free base 

C C N is soluble at 15** in 80 parts of water and at 100° in 

rr/^ r< f^f^TT ^5 P^rts. Codcinc differs from most of the other 

I 1 1 I I ' alkaloids, morphine, for example, in its relatively 

CHiO.C C C CHa high solubility in water. Alcohol, ether, amyl alcohol, 

\/\^\/ chloroform and benzene also dissolve codeine freely. 

I I I ^^ ^* however, practically insoluble in petroleum 

O— HC CH ether. Aqueous codeine solutions are strongly alka- 

\/ line and bitter. Pure codeine does not reduce iodic 

C acid, nor does it immediately produce a blue color or 

_j \ a blue precipitate in a mixture of potassium ferri- 

cyanide and ferric chloride solutions. A pure 

codeine solution is also not colored blue by ferric chloride solution alone. 

(Difference between morphine and codeine.) Phospho-molybdic acid, iodo- 

potassium iodide, potassium bismuthous iodide and potassium mercuric iodide 

give precipitates even with very dilute codeine solutions. On the other hand 

tannic and picric acids, gold and platinum chlorides are less sensitive. 

Detection of Codeine 

I. Sulphuric Acid Test. — Concentrated sulphuric acid dis- 
solves codeine without color. After long contact or upon appli- 



NON-VOLATILE POISONS 107 

cation of gentle heat, the solution will have a reddish to bluish 
violet color. The solution of codeine in concentrated sulphuric 
acid, heated to about 150° and then cooled, is colored deep red 
by a drop of concentrated nitric acid. 

2. Nitric Acid Test. — Cold nitric acid (25 per cent.) will 
convert codeine into nitro-codeine (Ci8H2o(N02)N08). At the 
same time the acid will dissolve the alkaloid with a yellow color 
which soon changes to red. Concentrated nitric acid dissolves 
codeine with a reddish brown color. 

3. Oxidation Test. — Mix a little codeine upon a watch glass 
with four times the quantity of finely powdered potassium arsen- 
ate (KH2ASO4). Add a few drops of concentrated sulphuric 
acid and then warm gently over a small flame. The acid will 
have a deep blue or blue-violet color, if the codeine is not quite 
pure. Excess of potassium arsenate does not aflfect the test. 
If water or sodium hydroxide solution is added, the blue color 
will change to orange-yellow. 

A trace of ferric chloride solution may be substituted for potassium arsenate. 
Sulphuric acid containing i drop of ferric chloride solution to 10 cc. of add is 
prescribed by the German Pharmacopoeia for detecting the alkaloid in codeine 
phosphate. 

4. Froehde*s Test. — This reagent dissolves codeine with a 
yellowish color which seon changes to green and finally to blue. 
Gentle warming of the solution over a very small flame will 
hasten this change of color. 

R. Mauch warms 2 or 3 drops of a chloral hydrate solution of codeine with i 
drop of Froehde's reagent. An intense blue color finally appears. 

5. Formalin-Sulphuric Acid Test.^ — Concentrated sulphuric 
add containing formalin dissolves codeine with a reddish violet 
color which changes to blue-violet. This color is persistent. 
The spectrum shows an absorption of orange and yellow. 

6. Furfurol Test.^ — Dissolve codeine in a few drops of con- 

* See preparation of reagents, page 314. 

• This test depends upon furfurol formed by the action of concentrated sul- 
phuric acid upon cane sugar. Very dilute aqueous furfurol solution (1 : 1000) 
may be substituted for cane-sugar. Excess of furfurol unlike cane-sugar does 
not interfere with the test. Tr. 



108 DETECTION OF POISONS 

centrated sulphuric add and warm very gently with a drop of 
cane-sugar solution which must not be in excess. This will 
produce a purple-red color. 

This test may also be made by mixing a drop of sugar solution 
with codeine, dissolved in about 5 drops of 50-60 per cent, aque- 
ous chloral hydrate solution, and then adding 1-2 cc. of con- 
centrated sulphuric add as an underlayer. A carmine red 
ring will appear at the zone of contact. The color is quite 
permanent and increases in intensity upon standing. If the 
sulphuric add and chloral hydrate solution are thoroughly 
mixed, the entire liquid will be red. After a time the shade of 
color will be more of a red-brown. 

7. Pellagrins Test. — Both codeine and morphine give this 
test. Dissolve codeine in concentrated hydrochloric add and 
add at the same time 3-4 drops of concentrated sulphuric add. 
Expel hydrochloric acid upon the water-bath and heat the resi- 
due about 15 minutes. Dissolve the dirty red or violet residue 
in 2-3 cc. of water, add a few drops of hydrochloric add and 
neutralize with acid sodium carbonate. Then add alcoholic 
solution of iodine drop by drop (2 to 4 drops) and shake thor- 
oughly for several minutes. An emerald green solution indi- 
cates codeine. Extract the green solution with ether. The 
color of the ether will be red, whereas that of the aqueous solu- 
tion will remain green. This is a test for apomorphine (see 
page 123) formed from codeine by the mineral add. 

Ci7Hi,(CH,)N0, + HCl = CitHitNO, + CH«.C1 + H,0 

Codeine Apomorphine 

8. Mecke's Test — The reagent, consisting of selenious add 
and concentrated sulphuric add,^ dissolves codeine with a blue 
color quickly changing to emerald green and finally becoming 
a permanent oUve green. 

NARCOTINE 

Narcotine, C22H2SNO7, crystallizes in shining prisms or in tufts of needles 
which are nearly insoluble in cold water but readily soluble in boiling alcohol 
or chloroform. Separation of alkaloid from the cold alcoholic solution is almost 

* See preparation of reagents, page 315. 



Y 

HC— 



NON-VOLATILE POISONS 109 

complete. At 15° narcotine dissolves in 170 parts of ether; 31 parts of acetic 
ether; and 32 parts of benzene. Solutions of narcotine are not alkaline nor 

bitter. In these respects narcotine is very 

OCH| different from the other opium alkaloids. Salts 

1 of narcotine do not crystallize, their stability is 

^x slight and their solutions react add. Salts with 

HC C.OCHi volatile adds are decomposed, when their solutions 

I jl are evaporated, with separation of narcotine. So- 

"^ C.CO dium acetate predpitates free narcotine from its 

solution in hydrochloric add. 

Constitution. — Narcotine is a monadd, tertiary 

base and as such combines with i mol. of CHi.I, 

CU O r rw forming narcotine methyl iodide (CtiHtiNOr.- 

* ^\y\ CHiI). This compound is formed at ordinary 

yO — C C N.CHi temperatures but the reaction is hastened by 

HsC^ I II I heat. Narcotine heated with hydriodic add loses 

^^^~^ ^ ^H« 3 methyl groups which form CHJ. The al- 

^^ ^^ akloid must therefore contain 3 'methoxyls, 

H Hi ' 3(CH«0-) groups, in the molecule. Heated with 

water to 140*', with dilute sulphuric add, or even 
with barium hydroxide solution narcotine is hydrolyzed into nitrogen-free 
opianic add and into the basic and consequently nitrogenous hydrocotamine: 

CnHtJ^OT + HtO = CioHioO, + Ci.HiJ^O, 
Narcotine. Opianic Hydrocotarnine. 

acid. 

By oxidative deavage, that is, by treatment of narcotine with such oxidizing 
agents as nitric add, manganese dioxide and sulphuric add, lead dioxide and 
ferric chloride, cotamine and opianic add are the products: 

CtJiiJ^GT + (H,0 + O) = CioHioO. + Ci,HisN04 
Narcotine Opianic Cotamine 

acid 

Evidently these deavage products show that this alkaloid is made up of two 
complexes, one nitrogen-free and the other containing nitrogen. The chemical 
constitution of these deavage products has been determined and is expressed by 
the following formulae: 

H CH,0 

C I H, 

^\ C C 

CH,0— C CH ^\/\ 

I II /O— C C N— CH, 
CHK)— C C— C=0 H,C< I [I I 

\/ H Nd—C C CH, 

c c 

HO— C=0 H H, 

Opianic Acid Hydrocotamine 



110 DETECTION OF POISONS 



CH,0 O 

C C— H 

xO— C C NH.CH, 
H,C< I 



O— C C CH, 

c c 

H Hi 

Cotamine 

On the basis of these results, Roser and Freund havie proposed the structural 
formula for narcotine given above. They consider the constitution of this alka- 
loid as definitely settled. If the formula of narcotine is compared with that of 
hydrastine (see page 112), a great similarity in structure will be seen. In fact 
narcotine is a methoxylized hydrastine. 

Detection of Narcotine 

Narcotine is so feebly basic that chloroform will extract the 
alkaloid completely from an aqueous tartaric acid solution. 
Consequently its separation from the rest of the opium alkaloids 
as well as from other alkaloids is easy. Naturally ether or 
chloroform will also extract narcotine from an aqueous alkaline 
solution. The alkaloid as it comes from its ether solution is 
usually a slightly colored, varnish-like residue which hardens 
after a time to a mass of radiating crystals. Narcotine is 
precipitated from its hydrochloric or sulphuric acid solution 
by iodo-potassium iodide, phospho-molybdic acid, potassium 
mercuric iodide, potassium bismuthous iodide even in consider- 
able dilution (i :5ooo). 

1. Sulphuric Acid Test. — -Dissolved with stirring in concen- 
trated sulphuric acid, narcotine produces a greenish yellow color 
which gradually changes to reddish yellow and finally after 
several days to raspberry red. 

2. Dilute Sulphuric Acid Test. — A solution of narcotine in 
dilute sulphuric acid (i 15), evaporated on the water-bath in a 
porcelain dish or over a very small flame, has a reddish yellow 
color, changing with stronger heat to crimson red. As the acid 
begins to evaporate, blue-violet streaks radiate from the margin 
and finally the entire liquid has a dirty red violet color (Dragen- 
dorff's reaction). The same color changes appear, if the yellow- 



NON-VOLATILE POISONS 111 

ish solution of narcotine in concentxated sulphuric acid is 
heated very carefully. 

3. Ftoehde's Test — This reagent dissolves narcotine with a 
greenish color. K concentrated Froehde's reagent is used, the 
green color changes immediately to cherry red, especially upon 
application of gentle heat. This color is quite persistent. 

4. Couerbe's Test — Dissolve narcotine in cold concentrated 
sulphuric add and mix a trace of nitric acid with this solution 
after 1-2 hours. A red color will appear and gradually become 
more and more pronounced. 

Erdmann's reagent gives the same color change. 

5. Wangerin's Test^ — Place a mixture of o.oi gram of 
narcotine with 20 drops of pure concentrated sulphuric acid 
and 1-2 drops of i per cent, cane sugar solution upon a watch 
glass and heat upon the water-bath with stirring about i minute. 
At first the solution has a greenish yellow color which passes 
through yellow, brownish yellow, brown and brown-violet into 
an intense blue-violet. 

The intensity of this color increases somewhat upon standing 
and the blue-violet color persists several hours. 

Applied to apomorphine, atropine, brucine, quinine, codeine, caffeine, hydras- 
tine, morphine, physostigmine, pilocarpine and strychnine, this test gives 
solutions that are colorless or nearly so. Only the morphine solution after a 
while has a pale pink color. Coniine and narcotine have a light yellow color; 
narceine chestnut-brown; and picrotoxin salmon color to pale pink. 

Colchicin, digitalin and veratrine behave toward this reagent as toward pure 
concentrated sulphuric acid without the addition of the small quantity of sugar. 

In this test 1-2 drops of i per cent, aqueous furfurol solution may be substi- 
tuted for the sugar solution. From yellow, brown, olive and other colors there 
finally emerges a deep, clear, dark blue. The brilliancy of this color increases 
somewhat on standing. After several hours there is a gradual change to a pure 
green color. For the detection of traces of narcotine (o.ooi gram) use a i per 
cent, sugar solution. 

6. Selenious Acid-Sulphuric Acid Test. — This reagent dis- 
solves narcotine with a greenish steel-blue color which after a 
time becomes cherry-red. Heat immediately discharges the 
cherry-red color. 

* Pharmazeutische Zeitung, 48, 607 (1903). 



112 DETECTION OF POISONS 

HYDRASTINE 

Hydrastine, C21H21NO6, occurs together with berberine, CmHitN04, and cana- 
dine, C20H21NO4, in hydrastis root, the root of Hydrastis canadensis, to the 

amount of 1.5 per cent, and more. The fluid ex- 

O.CH« tract prepared from this root and used in medicine 

J, contains 2-2.5 pcr cent, of hydrastine. 

^\ Preparation. — Extract hydrastis root with hot 

HC C.OCHs water containing acetic add. Filter the solution, 

I \\ evaporate to a thin extract and add 3 vols, of dilute 

^ <-.v.^ sulphuric acid (i : 5). Nearly all the berberine 

Q ^ separates out in fine yellow crystab as add sul- 

I phate, CsoHi7N04.H»S04. Predpitate hydrastine 

HC — O from the mother-liquor of berberine sulphate by 

HC CH means of ammonium hydroxide solution and purify 

/^\/\ the alkaloid by crystallization from acetic ether or 

yO—C C N.CH« alcohol. Hydrastine crystallizes from alcohol in 

HaCv I 'I I rhombic prisms melting at 132®. It is nearly in- 

""^ y^ ^"* soluble in water but freely soluble in hot alcohol, 

Q c benzene or chloroform. This alkaloid has a bitter 

H Hs taste and its solutions are alkaline. Hydrastine 

solutions are optically active. In chloroform this 
alkaloid is Isvo-rotatory, whereas in dilute hydrochloric add it is dextro- 
rotatory. 

Constitution. — The constitution of hydrastine is entirely analogous to that of 
narcotine (see page 109). On oxidation with dilute nitric add hydrastine gives 
opianic add and hydrastinine: 

C«H«NO. + (H,0 + O) = CoHioOs + CnHuNOa 
Hydrdstine Opianic acid Hydrastinine 

Hydrastine is a monadd base which is shown to be a tertiary base by its 
behavior toward alkyl iodides, for example, with CHiI it forms hydrastine methyl 
iodide, CaiHtiNOe.CHjI, which crystallizes in needles. Hydrastine contains 
two methoxyl groups, because when heated with hydriodic add according to 
Zeisel's method two such groups are removed. 

Since the chemical nature of opianic add has long been known, the only 
problem is the explanation of the nature of hydrastinine, the other cleavage 
product. The constitution of hydrastinine, as well as that of many other alka- 
loids, has been determined by A. W. Hofmann's method of exhaustive methyla- 
tion.^ Hydrastinine (I) is a secondary base which forms, when heated with an 

^ When the nitrogen of an organic base becomes quinquevalent, it is more sub- 
ject to change. Hofman (Liebig's Annalen, 78, 263 (1851) showed, for example, 
that tetra-ethyl-ammonium hydroxide breaks up on heating into triethylamine, 
ethylene and water: 

C2H1 



^*2*>N-0H = II + ChAn + H2O. 



Nitrogen in alkaloids on treatment with an alkyl haloid {e.g., CH«I) combines 
with it in many instances, forming compounds having a structure analogous 



NON-VOLATILE POISONS 113 

excess of CH|I, hydrastinine hydriodide and trimethyl-hydrastyl-ammonium 
iodide (II). Heated with alkalies, this ammonium iodide is decomposed into 
trimethylamine, hydriodic add and nitrogen-free hydrastal (III). The latter 
on oxidation gives hydrastic add (IV) which was recognized as the methylene 
ether of nor-meta-hemipinic add (V) : 

yCHlO + 2CH,I = 

a) (CH,0,)C,H< 

\CH,.CH,.NH.CH, 

/CH:0 
(CH,Ot)C»H< V 

\CH,.CH,.N(CH,)«I 

Hydrastinine Trimethyl-hydrastyl- 

ammonium iodide 
yCKlO 

(11) (CH,0,)C,H< 

\CH,.CH,.N(CH,) J + KOH = 

yCKlO 

(CH,0,)C»H,< + KI + HtO + N(CH,), 

^CH:CH, 
Hydrastal 
CH * O COOH 

(HI) (CH,0,)C»Ht<f Oxidized = (CH,0,)CJI,/ (IV) 

^CHiCH, ^COOH 

« Hydrastic acid 

Hydrastic add and nor-meta-hemipinic add are identical. The latter has the 

structure (V): 

H 
C 

^\ 
/O— C C.COOH 

(V) H,C< T II 

\)—C C.COOH 

c 

H 

Nor-meta-hemipinic acid 

From these and other relations it has been determined that cotarnine is a 

methoxy-hydrastinine : 

H CH,.0 H 

H / II 

C C=0 C C=0 

CC C NH.CH, yO— C C NH.CH, 

C C CH, ' S>- C C CH, 

c c c c 

H H, H H, 

Hydrastinine Cotarnine 

The alkaloid narcotine is a methoxy-hydrastine (see page zog). 

to that of tetra-ethyl-ammonium hydroxide. This process is called "exhaustive 
methylation." Upon decomposition these derivatives yield products which 
often throw light upon the structure of the alkaloid. 
8 



114 DETECTION OF POISONS 

Detection of Hydrastine 

1. Concentrated Sulphuric Acid dissolves hydrastine without 
color but upon being gently warmed the solution becomes violet. 

2. Froehde's Reagent dissolves hydrastine with a green color 
which gradually changes to brown. 

3. Mandelin's Reagent dissolves hydrastine with a rose- 
red color which immediately changes to orange-red and gradu- 
ally fades. 

4. Fluorescence Test. — Dissolve hydrastine in dilute sul- 
phuric acid, shake vigorously and add drop by drop very 
dilute potassium permanganate solution. Hydrastinine is 
formed and the solution shows a beautiful blue fluorescence. 

The ether extract of the alkaline solution on evaporation 
leaves hydrastine in a crystalline condition. 

QummE • 

Quinine, CsoH24N202, is precipitated amorphous and anhydrous from solutions 

of its salts by caustic alkalies, alkaline carbonates or ammonia. On standing, 

l£ however, it gradually becomes crystalline, forming a 

C hydrate with 3 molecules of water of hydration. 

/ \ * There are also other hydrates of quinine. Anhy- 

"*^ i CH.CHiCHj drous quinine melts at 173**; the trihydrate at 57°. 

I ^ An ether solution on evaporation usually deposits 

HO.C CHjCHj this alkaloid as a resinous, or varnish-like, amorphous 

\ I / residue. Quinine is soluble in about 2000 parts of 

I cold and 700 parts of boiling water; and freely solu- 

QH ^^^ ^" alcohol, ether or chloroform. Solutions of 

I H quinine in sulphuric, acetic or tartaric acid exhibit a 

C C beautiful blue fluorescence. In the case of the sul- 

r )< >, r\r^rT phatc this fluorescence is distinctly visible in a dilu- 
I 11 I tion of I : 100,000. 

HC C CH Hydrochloric, hydrobromic and hydriodic add do 

\/\y^ not give fluorescent solutions of quinine. These acids 

-^ ^ even discharge the fluorescence, if added to a fluor- 

escent quinine solution. 
Constitution. — Quinine is a diacid, ditertiar>' base, the salts of which with 
I and 2 equivalents of acid are usually crystalline. The salts with i equivalent 
of acid are the more stable. Quinine hydrochloride, C20H24N2O8.HCI.2H1O, 
used in medicine, crystallizes in long delicate tufts of needles. The diter- 
tiary character of quinine is shown by the fact that it unites with 2 mole- 
cules of methyl iodide, for example, to form quinine dimethyliodide, 
C2oH24N20j.2CH»I. Quinine must contain an hydroxyl group, since it can 
form a mono-benzoyl and a mon-acetyl-quinine. Moreover one methozyl 



NON-VOLATILE POISONS 



116 



group has been found in the quinine molecule. The difference empirically 
between dnchonine, Ci9H,,N,0, and quinine, C,oH,4N,0,, is CH,0. Every 
investigation of these substances has shown that quinine is a methoxy-dncho- 
nine. For example, on oxidation with chromic acid, dnchonine gives dnchonic 
add which was recognized as quinoline 7-carboxylic add; whereas quinine imder 
the same conditions gives quinic add, or p-methoxy-dnchonic add: 



C00H(7) 

HC C CH 

I i I 
HC C CH 

YY 

H 

Cinchonic acid 



C00H(7) 

(p)CH,O.C C CH 

I il I 
HC C CH 

\/\/ 
C N 

H 

Quinic acid 



Both alkaloids on oxidation also give the nitrogenous compounds mero- 
quinene, dnchoioijsonic acid and loijsonic add. Consequently there is no 
doubt that dnchonine and quinine contain two nitrogenous nucld, one of which 
is a quinoline complex. The second nudeus is connected with the latter in the 
7-position, as the formation of cinchonic and quinic adds shows. Meroquinene, 
dncholoiponic add and loijsonic add, derived by oxidation with chromic add 
from the so-called "second half'' of the dnchonine and quinine molecules, form 
a continuous series of oxidation products, since meroquinene can be oxidized to 
cincholoii>onic add and the latter to loijsonic add. The following formulae best 
explain the chemical behavior of these three comjsoimds: 



CH,.COOH 

1 




CH,.COOH 


COOH 


in 






CH 


CH 


H,C CH- 


-CH: 


CH, 


/\ 
H,C CH.COOH 


H,C CH.COOH 


H,C CH, 






1 1 
H,C CH, 


H,C CH, • 


\/ 






\/ 


\/ 


N 






N 


N 


H 






H 


H 


Meroquinene 




Cincholoiponic acid 


Loiponic acid 



The structural formula already given for quinine was proposed by W. Koenigs^ 
and is based on the results of his own experiments as well as on those of W. V. 
Miller and of Skraup. Cinchonine has hydrogen in place of the methoxyl group 
in the quinoline nudeus; otherwise the two alkaloids are identical in structure. 



Detection of Quinine 

Ether, benzene or chloroform will extract quinine from an 
aqueous alkaline solution. Ether on evaporation deposits the 

' Meroquinene and the Structure of the Cinchona Alkaloids; Annalen der 
Chemie und Pharmazie 347, 147 (1906). 



116 DETECTION OF POISONS 

alkaloid as a resinous, amorphous varnish in which its presence 
may be recognized by the following tests: 

1. Fluorescence Test — Dissolve the residue from the ether 
extraction of the alkaline solution in a little dilute sulphuric 
acid. If quinine is present, this solution will exhibit blue 
fluorescence. 

2. Thalleioquin Test. — Dissolve quinine in a few drops of 
very dilute acetic acid and add 5-10 drops of saturated chlorine 
water. The colorless solution has a faint, blue fluorescence. 
Excess of ammonium hydroxide solution will produce an emer- 
ald green color. A solution containing considerable quinine 
will give a green precipitate. This precipitate (thalleioquin) 
is always an amorphous substance, the composition of which 
has not been determined. It is soluble in alcohol and chloro- 
form but not in ether. 

E. Polacci recommends the following procedure for the thal- 
leioquin test. Gradually heat quinine (about o.oi gram) to 
boiling with a little lead dioxide (Pb02), 2-3 cc. of water and 2 
drops of dilute sulphuric acid. Let the solution settle and 
either decant or filter. Finally, carefully add 5-6 drops of 
ammonium hydroxide solution as a top layer. A fine green 
ring will appear at the zone of contact. 

Interferences with the Thalleioquin Test — Antipyrine interferes with this test. 
Mixtures of i per cent, solutions of antipyrine and quinine give finally a beautiful 
red instead of a green color. This interference does not cease until these two 
substances are in the proportion of 0.25 parts of antipyrine to 5 parts of quinine. 
Caffeine also interferes with the thalleioquin test, when the projsortion is 2 parts 
of quinine to 3 parts of caffeine. Other compounds like urea prevent the appear- 
ance of this color, whereas morphine, pilocarpine, cocaine, atropine, codeine, 
strychnine, carbolic acid and chloral hydrate have no effect upon the thalleioquin 
test. 

H. FUhner^ has shown that the thalleioquin reaction is connected with the 
p-oxyquinoline^complex. Chlorine passed into a solution of pure p-oxy-quino- 
line cooled with ice produces a white crystalline precipitate. This substance 
crystallizes from petroleum ether in colorless prisms or tabular crystals melting at 
58®. Structurally it is s,s-dichloro-6-keto-quinoline. Solutions of this dichloro- 
keto-quinoline and of its hydrochloride are colored a pure green or blue by am- 
monium hydroxide. Fuhner thinks s,6-quinoline quinone is probably formed 
and gives the green color with ammonia. 

* Berichte der Deutschen chemischen Gesellschaft 38, 2713 (1905). 



NON-VOLATILE POISONS 117 

H H H CI, HO 

C C C C C C 

HC C C.OH(p) HC C CO HC C CO 

HC C CH "" HC C CH ""hC C CH 

V\/ \/\/ \/\^ 

N C N C N C 

H H H 

p-Ozy-qainoline s.s-Dichloro-keto- 5,6-Qtiinoline 

quinoline qtiinone 

3. Herapathite Test. — Mix 30 drops of acetic acid, 20 drops 
of absolute alcohol and i drop of dilute sulphuric acid. Add 
20 drops of this mixture to o.oi gram of quinine and heat to 
boiling. Finally add i drop of an alcoholic solution of iodine 
(i : 10) or 2 drops of o.i n-iodine solution. When the solution 
has stood for some time, green leaflets with a metallic luster will 
form. This is an iodine compound of quinine called "Hera- 
pathite," having the constant composition 

4C20H24N2O2.3H2SO4.2HI.3H2O. 

This substance can be recrystallized from boiling alcohol. 
Herapathite crystals are pale olive-green by transmitted 
light but by reflected light they have a beautiful, cantharidin- 
green, metallic luster. 

Caustic alkalies, ammonia, sulphurous add and hydrogen sulphide decompose 
herapathite. A. Christensen recommends keeping on hand the following reagent 
for the herapathite test: 

Parts 
Iodine i 

Hydriodic acid (50%) i 

Sulphuric add o .8 

Alcohol (70%) so 

Add a few drops of this reagent to the alcoholic solution to be tested for quinine. 

4. Hirschsohn's Test.^ — If i drop each of 2 per cent, hydro- 
gen dioxide and 10 per cent, copper sulphate solution are added 
to a neutral solution of quinine hydrochloride or sulphate at 
boiling temperature, a more or less intense raspberry red color 
will appear. This color soon passes through blue-violet into 
blue and after a time into green. A quinine solution (i : 10,000) 
will still give a distinct red-violet color. 

^ Pharmazeutische Zentral-Halle 43, 367 (1902). 



118 DETECTION OF POISONS 

Excess of acid as well as of alcohol interferes with this test. The behavior 
of a solution of aloes toward this test is similar to that of quinine. 

Of the alkaloidal reagents potassium bismuthous iodide is 
especially recommended as a precipitant of quinine. With 
quinine sulphate solutions this reagent produces precipitates 
having an intense yellowish red color. Shaken with sodium 
hydroxide solution this precipitate is decomposed and unaltered 
quinine can be obtained by extraction with ether and evaporation 
of the ether solution. H. Thoms^ has made use of this reac- 
tion in the quantitative separation of quinine from mixtures. 

CAFFEINE 

Since caffeine (see page 79) is a weak base, ether will extract 
only a little of the alkaloid from the tartaric acid solution. The 
greater part will be in the ether extract of the alkaline solution. 
Ether usually deposits caffeine in white, shining needles ar- 
ranged in clusters. Caffeine dissolves in ether with some 
difficulty and the alkaline solution should be extracted several 
times. For the tests characteristic of this alkaloid see page 80. 

ANTIPYRINE 

Most of the antipyrine (see pag6 78) is obtained by extract- 
ing the alkaline solution with ether. It is usually purer from 
the acid than from the alkaline solution and frequently appears 
in crystalline leaflets. Antipyrine differs from most alkaloids 
in having only a faintly bitter taste and in being freely soluble 
in water. To identify antipyrine, dissolve the ether residue in a 
little water and divide the solution into two equal parts. Test 
one portion with ferric chloride solution and the other with 
fuming nitric acid. 

Detection of Antipyrine in Urine. — The color of urine after administration of 
antipyrine is intensely yellow to blood red. Part of the antipyrine in the organ- 
ism appears in the urine as oxy-antipyrine-glycuronic acid and another part is 
unchanged and can usually be detected directly in urine by ferric chloride solu- 
tion. A safer procedure is to add excess of ammonia to a considerable quantity of 
urine and extract with chloroform. Evaporate the solvent, dissolve the residue 

* Berichte der Deutschen pharmazeutischen Gesellschaft 16, 130 (1906). 



NON-VOLATILE POISONS 119 

in a little water and test the filtered solution for antipyrine with ferric chloride 
solution and with fuming nitric add. 

Antipyrine is easily absorbed. The urine may show a reddish color even an 
hour after the drug has been taken and give a test with ferric chloride solution. 
The red color disappears in about 24 hours but the elimination of antipyrine is 
not complete in that time. Its detection is still possible after 36 hours. A con- 
venient procedure is to add to the urine as an upper layer very dilute ferric chlo- 
ride solution. A red ring will appear if the urine contains antipyrine. Jonescu* 
states that antipyrine in the human organism passes imchanged into the urine. 
Only a small i>ortion — and large doses of the drug must have been taken — is 
eliminated in conjugation with sulphuric add. Conjugation with glycuronic 
add' (see above) according to Jonescu does not occur in the human organism. 

PYRAMIDONE 

Pyramidone, or 4-dimethyl-amino-antipyrine, CnHnNiO, has been exten- 
sively used in medidne of late as an antipyretic and anodyne. It is a white, 

crystalline powder, nearly tasteless and readily soluble 
* * in water. It melts at 108**. Its aqueous solution has 



N a neutral reaction. Ether removes only traces of 

/*\ pyramidone from add solution, but extracts it easily 

CHj Ni *C0 and completely from alkaline solution. Ether usually 

Q^ C»=^*C N(CH ) deposits this substance in fine needles. Pyramidone 

is also freely soluble in alcohol, ether, chloroform or 
benzene. It is a strong redudng agent and in this respect differs from antipyrine. 
For example, pyramidone will reduce gold chloride even in the cold, whereas 
antipyrine and tolypyrine require heat. 

Preparation. — Antipyrine dissolved in concentrated acetic add is converted by 
treatment with potassium nitrite into nitroso-antipyrine which appears as green 
crystals. This compound dissolved in alcohol may be reduced by zinc and acetic 
add to amino-antipyrine. The latter, dissolved in methyl alcohol and treated 
with methyl iodide and potassium hydroxide, is converted into dimethylamino- 
antipyrine, or pyramidone. 

C«Hs C«Hs CeH. 



N N N • 

CH,.N CO CH,.N CO CH,.N CO+aCH,! ^ 

CH,.C=CH HC).N0"*CH,.C=C.N0 ^ ~* CH,.C =C.NH, 2KOH 

Antipyrine Nitroso-antipyrine Amino-antipynne 

* Berichte der Deutschen pharmazeutischen Gesellschaft 16, 133 (i9<>6)- 

« Glycuronic add, C^HioO; « ^C(CH.0H)4C00H, may be regarded as a 

derivative of glucose. Possibly it occurs in normal urine in small quantity as 
a conjugated add. After administration of various alcohols, aldehydes, J^^'*^^<^' 
phenols (chloral hydrate, camphor, phenol, thymol, menthol, borneol), there 
takes place in the animal organism— often after oxidation or reduction a con- 
jugation of these substances with glycuronic acid. 



120 DETECTION OF POISONS 

C.H* 



N 

/\ 
CH,.N CO 

1 I 
CH,.C=C.N(CH,), 

Pyramidone 

Behavior in the Orgazusm. — Human urine, if neutral or faintly add, usually 
has a bright purplish red color after administration of pyramidone. After stand- 
ing for some time it will dejsosit a sediment consisting of red needles sduble in 
ether or chloroform but especially in acetic ether. JaS€^ recognized this com- 
pound as rubazonic acid, a pryazolone derivative. Isolation of rubazonic add 
from urine may be brought about as follows. Addify fresh urine with hydro- 
chloric add and let it stand in an open dish. The add will appear as small red 
particles. Ferric chloride solution produces a blue -violet color in the add liquid 
filtered from rubazonic add. This filtrate contains most of the product formed 
from pyramidone in animal metabolism, namely, crystalline antq>yryl-urea 
melting at about 245^. 

C»Hs 

I 
N 

/\ 
CH,J^ CO 

I I 
CH,.C =C.NH.CO.NH, 

Antipyryl-iirea 



Detection of 

1. Ferric Chloride Test — Ferric chloride solution added to 
pyramidone produces a blue-violet color which soon changes to 
reddish violet and then disappears. 

2. Fuming Nitric Acid Test — ^A few drops of fuming nitric 
acid, added to a solution containing pyramidone, give a blue to 
blue-violet color. 

3. Bromine Water Test — This reagent imparts a grayish 
color to pyramidone solutions. With concentrated solutions 
it produces an inky color. 

4. Iodine Test — ^Tincture of iodine colors an aqueous pyra- 
midone solution blue. 

^Berichte der Deutschen chemischen Gesellschaft 34, 3737 (1901); and 35, 
3891 (1902). 



NON-VOLATILE POISONS 



C. Eztractioii of the Ammoniacal Solution with Ether end 
Chlorofonn 



I 



(a) Ether Extract. — Apomorphine and traces of morphine.' 

(/3) Chloroform Extract.^ — Morphine and narceine. (It may 
also contain antipyrine and caffeine.') 

The aqueous alkaline solution (see page 8i), separated 
from ether, must be tested further for the substances under 
a and p. 

Apomorphine may be recognized by the green color of the 
aqueous acid solution. Excess of sodium hydroxide solution 
causes oxidation, especially if the solution is exposed for any 
length of time to air, and gradually changes the color to deep 
purple-red. Moreover, the ether extracts, both of the add 
and alkaline solutions, are red or violet-red when apomorphine 
is present. Solutions, examined by the Stas-Otto method, not 
having these characteristics, need not be tested for apomorphine. 
In that case proceed at once with the morphine and narceine 
tests. 

To extract apomorphine, morphine and narceine with the 
proper solvent, the aqueous solution separated from ether, which 
is alkaline from sodium hydroxide solution (see page 8i) , must 
be rendered alkaline with ammonium hydroxide solution. 
First acidify the solution with dilute hydrochloric add {test 
■with blue litmus paper) and then add ammonium hydroxide 
solution until alkaline. 

(a) If there is any indication of apomorphine, first extract the 
ammoniacal solution repeatedly with ether and then several 
times with hot chloroform for the morphine and narceine tests. 

((3) If there is no indication of apomorphine, extract the 
ammoniacal solution several times direct with hot chloroform 
(see below) . 



4 

i 



* Ether dissolves traces ot freshly precipitated, amorphous morphine. 

* Antipyrine and cafFeine, though freely soluble in chloroform, dissolve *ith 
difficulty in ether. The Utter solvent frequently falls to extract these suhstanceg 
completely from aqueous solution. They will then appear in the chloroform 
«ztrftct. 



122 DETECTION OF POISONS 



APOMORPHINE 



Constitution. — ^Apomorphine, CnHnNd, is a monadd, tertiary base with two 
phenol hydroxyl groups. According to R. Pschorr^ it has the structural formula 
here given. 

Properties. — ^Ajsomorphine is an amorphous base 

H H CH I'^dily soluble in alcohol, ether, benzene or chloro- 

C C N ^OTm and colored green in contact with air. Aqueous 

J'\/\/\ ^^^ alcoholic apomorphine solutions, originally color- 

HC C CH CHs less, soon turn green in the air from oxidation. Solu- 

I JJ 1 I tions of apomorphine thus changed by oxidation are 

(3)H0.C C^ C^ CHt emerald green. Ether and benzene solutions are 

C C C(8) purplish violet; those in chloroform blue- violet. Be- 

I II I ing phenolic in character, apomorphine resembles 

(4) HO HC CH morphine in its solubility in sodium hydroxide solu- 

\i tion. Alkaline solutions of the alkaloid absorb oxy- 

21 gen from the air and become brown or even black 

in color. AI>omorpi^ne differs from morphine in 

being more soluble in water and in alcohol, but especially in being soluble in 

ether, benzene and cold chloroform in which morphine is almost insoluble. 

Formation and Preparation. — Sulphuric, hydrochloric, phos- 
phoric and oxalic acids, the alkalies and zinc chloride have 
maitily a dehydrating action upon morphine and convert it 
into apomorphine: 

C17H1.NO, = HjG H- CitHitNOi 

Morphine Apomorphine 

Codeine, the methyl ether of morphine, also gives apomor- 
phine when heated at 140® with concentrated hydrochloric acid. 

CitHhNOjCOCH,) -h HCl = H2O -h CHiCl + CnHnNO, 

Codeine Apomorphine 

Apomorphine is prepared by heating morphine (i part) with 
concentrated hydrochloric acid (20 parts) for 3 hours in an 
autoclave at 130-150®. 

(a) Detection of Apomorphine in the Ether Extract 

Ether will not extract apomorphine from a solution contain- 
ing tartaric acid but will dissolve its colored oxidation products. 
This solvent behaves similarly toward solutions of this alkaloid 
in sodium or potassium hydroxide solutions. Ether or chloro- 

* Berichte der Deutschen chemischen Gesellschaft 39, 3124 (1906); and 40, 
1984 (1907). 



NON-VOLATILE POISONS 123 

form will extract apomorphine only from a solution alkaline 
with ammonium hydroxide. Ether solutions of apomorphine 
usually deposit a greenish residue. A characteristic of this 
alkaloid is its strong reducing action. For example, it will re- 
duce iodic acid with liberation of iodine and produce a purple 
color with gold chloride. Apomorphine gives the following 
tests: 

1. Sulphuric and Nitric Acids. — Concentrated sulphuric acid 
dissolves apomorphine without color. Addition of a drop of 
concentrated nitric acid to such a solution produces an evan- 
escent violet color that soon changes to blood red and finally to 
yellowish red. With concentrated nitric acid alone this alka- 
loid gives a violet-red color that soon becomes red-brown and 
finally brownish red. 

2. Pellagri's Test — Dissolve apomorphine in dilute hydro- 
chloric or sulphuric acid and first add acid sodium carbonate in 
excess. Then add drop by drop 1-3 drops of an alcoholic iodine 
solution and shake for several minutes. The solution will have 
a blue-green or emerald green color. Extract with a little ether 
and the solvent will become violet, whereas the aqueous solution 
will remain green. 

3. Froehde's Test. — This reagent dissolves pure apomorphine 
with a green color. If the alkaloid has been acted upon by air 
to any extent, the color is violet. 

4* Wangerin's^ Test. — Prepare a fresh solution of apomor- 
phine hydrochloride (about i per cent.). Add 4 drops of potas- 
sium dichromate solution (0.3 per cent.) to i cc. of this solution 
and shake for about i minute. The solution will have an in- 
tense dark green color. Then add 10 cc. of acetic ether and 
shake again. This solvent will become violet. Finally add 
from a pipette about 5 drops of stannous chloride solution^ 
(i per cent.) and shake well. The color of the acetic ether layer 
will change to green and, upon further addition of a few drops of 

* Pharmazeutische Zeitung 47, 599 and 739-740 (1902). 

' Prepare this reagent as follows: 

Crystallized stannous chloride (SnCls.aHsO) i gram 
Hydrochloric add (25 per cent.) 50 cc. 

Water 50 cc. 



124 DETECTION OF POISONS 

potassium dichromate solution, the acetic ether will again be- 
come violet. If lo cc. of chloroform are substituted for acetic 
ether in this test, the oxidation product of apomorphine will im- 
part the same violet color to the chloroform. But if stamious 
chloride solution is added carefully, the color will change to pure 
indigo blue and persist upon further agitation with potassium 
dichromate solution. 

5. E. Schmidt's Tests. ^ — (a) A drop of very dilute ferric 
chloride solution (i : 100) will color 10 cc. of an aqueous apo- 
morphine hydrochloride solution blue even in a dilution of 
I : 10,000. 

(ft) Shake 10 cc. of the same apomorphine hydrochloride 
solution with i cc. of chloroform. Then render alkaline with 
sodium hydroxide solution and at once shake with air. The 
aqueous solution becomes evanescent violet in color and the 
chloroform blue. 

(/?) Examination of the Chloroform Extract 

Preliminary Morphine Test. — As a preliminary test for mor- 
phine, acidify a small portion of the aqueous alkaline solution 
separated from ether (see page 81) with dilute sulphuric acid, 
add iodic acid solution and extract with a little chloroform. If 
the latter has a violet color from dissolved iodine, morphine 
may be present. But a final conclusion regarding the presence 
of morphine must not be drawn from a positive test, since there 
are many other organic substances besides this alkaloid that 
will reduce iodic acid.* This is a delicate preliminary test for 
morphine and that is its only value. If it is negative, morphine 
is probably absent. 

To detect morphine and narceine positively, render the aque- 
ous solution alkaline with ammonium hydroxide and extract at 
once as already directed (see page 121) with considerable hot 
chloroform' in a capacious flask. Separate the two liquids as 

» Aj)Otheker-Zeitung 23, 657 (1908). 

* In testing animal matter that contained no morphine, the author has repeat- 
edly obtained extracts that strongly reduced iodic add. 

»C. Kippenberger (Zeitschrift ftir analytische Chemie 39, 201, 290) uses 
chloroform, containing 10 per cent, of alcohol by volume, to extract morphine. 






NON-VOLATILE POISONS 125 



dsual in a separatory funnel- Several extractions of the aque- 
ous solution with fresh portions of hot chloroform are necessary 
because of the slight soiubiHty of morphine even in boiling 
chloroform. Should the chloroform and the aqueous solution 
form a refractory emulsion that will not separate, add a few 
drops of alcohol, set the flask on a warm but not boiling water- 
bath and carefully turn the Hask from time to time. This 
procedure usually causes the immediate separation of the two 
liquids. Place the combined chloroform extracts in a dry flask, 
add a few crystals of dry sodium chloride or anhydrous sodium 
sulphate to remove adherent water, pour the chloroform when 
clear through a dry filter and evaporate in not too large a glass 
dish placed upon a warm water-bath. The chloroform may also 
be filtered directly into the dish as fast as it evaporates. If the 
residue is bitter and can be scraped together with a platinum 
spatula or a pocket knife, test for morphine and narceine.' In 
testing for morphine use Froehde's, Husemann's and Pdlagri's 
tests as well as those given by formalin-sulphuric acid and iodic 
acid. The presence of morphine is not established unless all 
these morphine tests give positive results. If the quantity of 
the residue from chloroform permits, test for morphine with 
ferric chloride solution. This test is very characteristic of 
morphine but requires more than traces for a satisfactory result. 

PuriScatioii of In^nire Morphine 

When the chloroform residue is too impure, especially if red 
or brown, it must be purified. Dissolve in hot amyl alcohol 
and shake the solution thoroughly with several portions of hot 
water containing a few drops of dilute sulphuric acid. The acid 
dissolves the morphine, whereas the amyl alcohol retains most of 
the coloring matter. Add ammonium hydroxide solution in 
excess to the acid solution and extract several times with hot 
chloroform. The morphine obtained by evaporation of the 
chloroform should be nearly pure. 

' Andpynne Btid caffeine may also be in Ibis residue (see above). 



126 DETECTION OF POISONS 

MORPHINE 

Morphine, CirHisNGs, crystallizes from dilute alcohol in shining prisms 
which are colorless and transparent and but slightly soluble in water (1:5000 

at IS®; and 1:500 at 100®). These solutions are very 

TT TT I bitter and have an alkaline reaction. Crsrstalline mor- 

(2 (^ 2^ phine is insoluble in ether and benzene. The amor- 

^\/\/\ phous alkaloid is soluble in amyl alcohol, hot chloroform 

HC C CH CHi and acetic ether. Solutions of the hydroxides of ammo- 

HO C C c CH ^^* potassium or sodium and sodium carbonate solution 

* \/\/\/ precipitate free morphine from solutions of morphine 

C C CH salts. 

o— c CH, Constitution. — Morphine is a monacid, 

H C tertiary base whose nitrogen is in union 

/\ with three atoms of carbon. The three 

H OH 

oxygen atoms have different functions. One 
is a phenolic hydroxyl and gives to morphine the character of 
a monatomic phenol. Consequently when sodium hydroxide 
solution is added drop by drop to a morphine salt solution, 
there is first a precipitate of crystalline morphine (a) which is 
freely soluble in excess of alkali (jS) but is again precipitated on 
addition of ammonium chloride solution (7) : 

(a) CnH,8N0,(0H).HCl + NaOH = CirHigNOiCOH) + H,0 -f NaCl, 

(/3) Ci7H,gN02(OH) + NaOH = Ci7H,8NO,(ONa) + H,0, 

(v) Ci7H,8NO,(ONa) + (H4N)C1 = CnHisNOjCOH) + NH, -f NaCl. 

Hydrogen of this phenolic hydroxyl may be replaced also by 
alkyl groups and acid radicals. In codeine this hydrogen is 
replaced by methyl. A second oxygen atom of morphine is 
alcoholic and the third is indifferent. The latter like the oxygen 
of an ether is combined with two carbon atoms and forms a so- 
called bridge-oxygen atom. 

Of the 17 carbon atoms of morphine 14 belong to the phenan- 
threne nucleus,^ since the nitrogen-free cleavage products of 

CH=-CH CH=CH 

»Phenanthrene,Ci4Hio,HC;^ \c — c/ ^^^' occurs in 

CH C C CH 

CH=CH 
coal-tar together with anthracene. It forms colorless crystals which melt at 99' 
and boil at 340°. It is readily soluble in ether or benzene and with difficidty in 
alcohol. Phenanthrene solutions exhibit bluish fluorescence. 



NON-VOLATILE POISONS 127 

morphine and codeine, namely, morphol and morphenol, have 
been identified as phenanthrene derivatives. R. Pschorr has 
synthesized morphol which is 3,4-dioxyphenanthrene. Mor- 
phenol contains two hydrogen atoms less and may be converted 
into morphol by reduction with nascent hydrogen. These 
two phenanthrene derivatives have the following structural 
formulae: 



H 


H 


C 


C 


/\ 


/\ 


HC CH 


HC CH 


i in 


i i 


h/Y 


HC C \ 


H^ i 


1 
HC C / 


\/\ 


\/\/ 


C C.OH(4) 


c c 

HC C.OH 


(i)HC C.0H(3) 


<y 


C 


H 


H 


Morphol 


Morphenol 



By distillation over zinc dust morphenol may be reduced to 
phenanthrene. 

The structural formula of morphine written above was pro- 
posed by R. Pschorr^ and seems to explain most satisfactorily 
the reactions of this alkaloid. 

Morphine is easily oxidized. This may be brought about in 
alkaline solution by atmospheric oxygen. Potassium permanga- 
nate or ferricyanide and ammoniacal copper solution may also 
be used. As a result the non-toxic oxy-dimorphine, also called 
pseudomorphine, which is soluble in caustic alkali, is formed: 

2C,7H„NO, + O = (CnHisNO,), + H,0. 
M orphine Oxydimorphine 

Detection of Morphine 

I. Nitric Acid Test. — Concentrated nitric acid dissolves mor- 
phine with a blood red color which gradually changes to yellow. 
Stannous chloride or ammonium sulphide solution will not re- 

^ Berichte der Deutschen chemischen Gesellschaft 40, 1984 (1907). 



128 DETECTION OF POISONS 

Store the violet color of a solution that has become yellow. 
(Distinction from brucine.) 

2. Husemann's Test — ^Dissolve morphine upon a watch glass 
in a few cc. of concentrated sulphuric acid. The solution is col- 
orless. Heat for 30 minutes upon the water-bath, or over a 
small flame for a very short time until white fumes arise. A 
reddish or brownish color appears. Cool and add 1-2 drops of 
concentrated nitric acid. A fugitive, reddish violet color ap- 
pears and soon changes to blood red or yellowish red. This 
color gradually disappears. 

A preferable procedure is to dissolve morphine in cold con- 
centrated sulphuric acid and add a trace of concentrated nitric 
acid after the solution has stood in a desiccator 24 hours. A 
small crj-stal of potassium nitrate or chlorate may be substituted 
for nitric acid. 

Frequently impure morphine is obtained from the chloroform extrmct of a 
solution prepared from, animal material. Such a residue gives a more or less 
highly* colored solution with sulphuric acid. Heat usuaUy intensifies the color. 
But exTn under these conditions it is possible to detect the red color caused by 
nitric acid or potassium nitrate. 

3^ Pellagrins Test — Proceed as described for codeine. (See 
page 108.) Avoid excess of alcoholic iodine solution, otherwise 
the latter may mask the green color. 

4* IVoehde's Test — This reagent dissolves morphine with a 
violet color which passes through blue to dirty green and finally 
to faint red. Tht^e a^lors vanish on addition of water. 

5* Fomuddehyde-Su^uiic Add Test — The solution used 
for this tt^t is calltxl Marquis* reagent*. With a trace of mor- 
phine it prixluces a purple-red color which changes to violet 
and iin;vlly bt\x\n\t'« pure blue. This blue solution^ kept in a 
tt'^t-lubo and only slightly exixvMfd to air. retains its color for 
some time. l\Hleine aiui a^xunorphine give the same violet 
cv^lor. Narvvtino als^^ gives viixlet solutions but they become 
olive gnvu and unally yellow. Oxy-dimorphine gi>*es a green 
cv^lor. 

^ Mi\ t-^t \i(\^(vjt vxf 4x^ |v^r ct^nt. l\>rmaKklo\le sotutioQ vich 5 cc. o£ coDcm- 
tratiM »ul)>huric Acki ai\d usw^ a ifew dc\>|« ol ihb mixture for t&e mcxphiBe tcsL 



NON-VOLATILE POISONS 129 

6. Iodic Acid Test. — Shake a solution of morphine in dilute 
sulphuric add with a few drops of iodic acid and chloroform. 
Morphine will liberate iodine which will dissolve in chloroform 
with a violet color. 

Obviously this delicate test is conclusive for morphine only in the absence of 
other reducing substances. 

7. Ferric Chloride Test — Add 1-2 drops of neutral ferric 
chloride solution to a neutral solution of a morphine salt. A 
blue color appears. In testing the chloroform residue, dissolve 
in a little very dilute hydrochloric acid. Evaporate this solu- 
tion to dryness upon the water-bath, dissolve the residue in pure 
water and add a drop of ferric chloride solution. 

8. Lloyd's Test. — Lloyd has found that a mixture of morphine, 
hydrastine and concentrated sulphuric acid alone without 
potassium dichromate will produce the same violet color given 
by the latter with a solution of strychnine in concentrated sul- 
phuric acid. Lloyd's reaction is of value in the detection of mor- 
phine or hydrastine only when more than traces of both alka- 
loids are present. A. Wangerin^ considers these reactions 
characteristic only when 0.005-0.01 gram of morphine and 
0.002-0.01 gram of hydrastine are present. 

Make an intimate mixture of about these quantities of both 
alkaloids upon a watch glass. Add 5 drops of pure concen- 
trated sulphuric acid and stir the mixture for 10 minutes 
over a white background. In the center the color-tone is a 
clear red-violet and more or less of a blue-violet in the thinner 
marginal region. Apomorphine hydrochloride, treated in the 
same way with hydrastine and concentrated sulphuric acid, 
gives almost the same reaction as morphine. 

g. Prussian Blue Test. — Add a few drops of a dilute mixture 
of -ferric chloride and potassium ferricyanide solutions to a 
morphine salt solution. A deep blue color appears. Consid- 
erable morphine produces a precipitate of Prussian blue. 

Potassium ferricyanide oxidizes morphine to oxy-dimorphine: 

2Ci7H,»NO, -f 2KOH -f K6Fe,(CN)i2 = 2H2O -f (CnHisNO,), -f 2K4Fe(CN). 
Morphine Potassium Oxy-dimorphine Potassium 

ferricyanide ferrocyanide 

^ Pharmazeutische Zeitung 46, 57(1903). 
9 



NON-VOLATILE POISONS 131 

Behavior of Morphine m the Animal OrganisoL — ^The mucous lining of the 
stomach, rectum or respiratory passages as well as open wounds absorb mor- 
phine. The alkaloid injected hypodermically acts more rapidly and more po- 
tently than when absorbed from the stomach. Marquis^ found that morphine 
disappears very quickly from the blood but is firmly retained by certain organs 
like the brain. Some absorbed morphine is conjugated with glycuronic add and 
some is oxidized but the rest of the alkaloid is eliminated unchanged. Faust 
has found that morphine is transformed or destroyed only in men and animals 
habituated to the poison but is eliminated unchanged nearly quantitatively in 
the faeces in the case of organisms not immunized. Morphine appears in the urine 
only in very small quantity after medicinal doses. In men and dogs a not insig- 
nificant quantity of the morphine taken is eliminated by the glands of the gastro- 
intestinal tract, even when the alkaloid has been subcutaneously injected. 

Marquis found that more than 30 per cent, of intravenously injected 
morphine is deposited in the liver in the course of 15 minutes. The alkaloid is 
present at first In this organ in the free state and then is soon combined or trans- 
formed. The conjugation of morphine in the brain also begins very soon. Free 
morphine is also rapidly changed in the blood, spleen, kidne3rs and in the mucous 
lining of the intestines. Marquis states that always in acute and even more so 
in chronic morphine poisoning a large quantity of the poison leaves the blood and 
is stored in the salivary glands, mucous lining of the stomach and large intestine, 
kidneys, spleen, liver and is withdrawn by these organs from the brain and spinal 
cord. 

Morphine is quite resistant to putrefaction. The author' detected this alka- 
loid positively in animal material containing morphine (stomach and intestines 
together with contents) which had stood for 15 months in a glass vessel and had 
completely putrefied in presence of insufficient air. 

NARCEINE 

OCHi Narceine, Cj»Hj7NO|.3HjO, crystaUizes from 

C water or alcohol in prisms which melt at 165° 

^\ when air dried. The alkaloid has a faintly bitter 

^9 C.GCHs ^^^^ Though only slightly soluble in cold water, 

HC C.COOH *' ^ freely soluble in hot. When a hot saturated 

\/^ aqueous solution of narceine is cooled, it solidifies 

^ to a crystalline mass. Narceine is insoluble in 

1q ether, benzene or petroleum ether and is soluble 

QYLiO. I ®^y ^^ difficulty in cold alcohol, amyl alcohol 

C CHs or chloroform. In detecting narceine it is im- 

^\/ /CH3 portant to know that it is not extracted by ether, 

XT cc \ if I ^CH benzene or petroleum ether from a solution ren- 

^O.C C CHa dered alkaline by potassium or sodium hydroxide 

\/\/ solution. It is, however, extracted by hot chloro- 

C C form or amyl alcohol from an aqueous solution ren- 

* dered alkaline by ammonium hydroxide solution. 

'Arbeit en des Dorpater Instituts, ed Robert, 14 (1896). 

'Berichte der Deutschen Pharmazeutischen Gessellschaft 11, 494 (1901). 



132 DETECTION OF POISONS 

ConstitutioiL — Naxceine is a weak tertiary base in which two 
methyl groups are attached to nitrogen. By means of Zeisel's 
method it may be shown that the molecule also contains three 
methoxyl groups. Narceine, being soluble in caustic alkalies 
and forming esters with alcohols, must contain a carbozyl 
group. The alkaloid must also contain a carbonyl group (CO), 
since it forms a hydrazone with phenyl-hydrazine. The nar- 
ceine formula above may therefore be resolved into: 

CmHitNO,- CiiHnON (CH.), (OCH,), (CO) (COOH). 

The narceine molecule contains neither an alcoholic nor a 
phenolic hydrox>-l group, since it forms no acetyl derivative 
with acetic anhydride. There is a close relationship between 
narceine and narcotine. By heating narcotine iodo-methylate 
with sodium hydroxide solution Roser converted this compound 
into a base called pseudo-narceine. Freund has recently shown 
that Roser's pseudo-narceine is identical with the opium alka- 
loid narceine and explains the conversion of narcotine into nar- 
ceine bv sa\'in? that the iodo-methvlate loses i molecule of 
hydriodic add and takes up i molecule of water: 

OCH, OCH, 

i 
c c 

HC C-OCH, HC C.OCH, 

HC C.CO HC CCOOH 



\ 



/ 



c c 

-HrO 
CH4O H C— O = CHjO CO 

-HI 
C HC , C CHi 



/ 



\ -. I 



O.C C X-<H, O.C C X/ 

HtC \ CH; H^. XTH, 

O.C C CH: ^.C C CH, 

\ X ^ 

c c c c 

H H: H Hs 

All the reacdons and iransformations of narceine can easily 
be cxfJained on the basis of this strjcmrai formula. 



NON-VOLATILE POISONS 133 

Detection of Narceine 

1. Sulphuric Add Test. — Concentrated sulphuric add dis- 
solves narceine with a grayish brown color, which gradually 
changes to blood red. This reaction takes place at once with 
heat. 

2. Dilute Sulphuric Add Test — Narceine, warmed in a porce- 
lain dish upon the water-bath with dilute sulphuric acid until 
a certain concentration is reached, gives rise to a fine violet 
color which changes after long heating to cherry red. 

3. Froehde's Test — At first a solution of narceine in this 
reagent has a brownish green color which gradually changes 
to green and finally to red. Gentle heat hastens this reaction. 

4* Iodine Test — Aqueous iodine solution (iodine water) or 
iodine vapor colors solid narceine blue. 

Morphine interferes with or entirely prevents this reaction. 

5. Erdmann's Test — This reagent, as well as concentrated 
nitric add, dissolves narceine with a yellow color which heat 
changes to dark orange. 

6. Chlorine-Anmionia Test — Pour a few drops of chlorine 
water upon narceine and add, while stirring, a few drops of 
ammonium hydroxide solution. A deep red color immediately 
appears. 

7. Resorcinol-Sulphuric Acid Test*^ — Mix thoroughly upon 
a watch glass resorcinol (o.oi to 0.02 gram) with 10 drops of 
pure concentrated sulphuric acid. Add a trace of narceine 
(about 0.002 to 0.005 gram) and, while stirring, warm the^'m- 
tensely yellow solution upon a boiling water-bath. A carmine 
red to cherry red color appears. As the solution cools, this 
color begins at the margin to change gradually to more of a 
blood red and finally after several hours to orange-yellow. 

8. Tannin-Sulphuric Acid Test. — Mix narceine (0.002 to 
o.oi gram) with tannin (o.oi to 0.02 gram) and 10 drops of 
pure concentrated sulphuric acid. Heat with constant stirring 
upon the water-bath and the color of the solution, which is 
yellowish brown at first, soon becomes pure green. If heat is 

* A. Wangerin, Pharmaceutische Zeitung, 47, 916 (1902). 



134 DETECTION OF POISONS 

applied for some time, the green color changes to blue-green 
and finally through a more or less blue tone to a dirty green. 

Tannin-sulphuric add gives a similar color test with narcotine and hydras- 
tine which closely resemble narceine in constitution. 

Of the general alkaloidal reagents potassium zinc iodide^ 
precipitates narceine even in a dilution of i : looo. It is a 
white, filiform precipitate which after a time becomes blue. 
This blue color appears immediately, if a trace of iodine solu- 
tion is added to the reagent. 

Of the other general reagents iodo-potassium iodide, potassium mercuric 
iodide, potassium bismuthous iodide and phospho-molybdic add are characterized 
by considerable delicacy toward narceine. 

SYNOPSIS OF GROUP H 
Stas-Otto Method 

A. Ether Extract of Acid Solution may Contain : 

Picrotoxin. — Very bitter. Reduces Fehling's solution with 
heat. 

Melzer's test: red streaks radiating from picrotoxin with 
alcoholic benzaldehyde + cone. H2SO4. 

Cone. H2SO4: soluble with yellow or orange-red color; drop 
of K2Cr207 + Aq has brown margin. 

Langley's test: picrotoxin + 3 parts KNO3, moistened with 
cone. H2SO4, red with excess of saturated NaOH + Aq. 

Colchicin. — Very bitter. Yellowish and amorphous. Dilute 
mineral acids render aqueous solutions intensely yellow. 

Cone. HNO3: soluble with dirty violet color changing to 
brownish red and finally to yellow; excess of KOH + Aq 
renders orange-yellow or orange-red. 

Zeisel's test: boil yellow colchicin solution in cone. HCl in test- 
tube 2-3 minutes with 2 drops of FeCU+Aq. Green or olive- 
green when cold, especially if diluted with equal volume of water. 

Picric Acid. — Very bitter. Yellow. Material and extracts 
more or less intensely yellow. 

^ See page 312 for the preparation of this reagent. 



NON-VOLATILE POISONS 



135 



VtH-l 






IsopuTpuric acid test: aqueous picric acid, gently warmed 
with a few drops of saturated KCN + Aq, gives red color. 

Picraminic acid test: aqueous picric acid, warmed with few 
drops of (H4N)2S + Aq, becomes red. 

Dyeing test: aqueous picric acid dyes wool and silk intense 
yellow but not cotton. 

Acetanilide. — Faint, burning taste, 

Indophenol test; heat with a few cc. of cone. HCl and evap- 

■ate to about 20 drops. Cool, add aqueous phenol solution 
and then calcium h>-pochlorite solution drop by drop. Mix- 
ture, shaken with excess of ammonia, becomes dirty red to 
blue-violet and blue. 

Phenylisocyanide test: boil with KOH + Aq and then add 
a little chloroform. Odor of phenyhsocyanide. 

Isolation of anihne: boil several minutes with alcoholic KOH, 
dilute with water and extract with ether. Evaporation of 
solvent leaves oily drops of aniline. Dissolve in water and test 
with calcium hypochlorite. 

Phenacetine. — Tasteless. Gives indophenol but not phenyl- 
isocyanide test. 

Cone. HNOj: yellow color even cold. Dil. HNO3 dissolves 
with yellow or orange-yellow color, if heated. Yellow nitro- 
phenacetine crystalhzes as saturated solution cools. 

Salicylic Acid. — Sweet, acidulous, harsh taste. 

FeCIj -f Aq: aqueous solutions colored blue-violet; jf dilute, 

;ore of a red-violet. 

Millon's test: red color upon warming. 

Brj + Aq: yellowish white, crystaUine precipitate. 

Veronal.— Bitter. Crystalline. 

Dissolve ether residue in very little NaOH + Aq or (H^N)- 
OH -I- Aq, filter and acidify filtrate with dil. HCl. Veronal 
crystallizes. Wash with a little cold water, dry and determine 
melting point (187-188°). The crystals mixed with pure 
veronal should have same melting point. 

Antipyrine. — Mild, bitter taste. Examine aqueous solution 
of ether residue for antipyrine. 

FeClj + Aq: red color. 



136 DETECTION OF POISONS 

HNOs: green color with 1-2 drops of fuming add. Heat 
and a few more drops of fuming add change green color to red. 

Most of the antipyrine in ether extract of alkaline solution 
(see B) . 

Caffeine. — Faintly bitter. 

Clj + Aq : evaporated upon water-bath with saturated 
CI2 + Aq, gives red-brown residue which turns purplish red 
moistened with very little (H4N)0H + Aq. 

Most of the caffeine in ether extract of alkaline solution 
(see B). 

Cantharidin.^ — Rhombic leaflets from ether solution. 

Physiological test: triturate residue with few drops of almond 
oil and test mixture as vesicant by applying to upper part of 
the arm. 

B. Ether Extract of Alkaline Solution may Contain : 

Coniine. — Yellow oil drops with penetrating odor. 

Cold saturated aqueous solution becomes milky when warmed. 

Spontaneous evaporation with a drop of HCl gives coniine 
hydrochloride as doubly refractive crystals which are needle 
or prism shaped and sometimes in star-like clusters. 

Physiological test: paralysis of peripheral nerves. 

Nicotine. — ^Liquid. Remains dissolved in residual water 
upon evaporation of ether and has faint tobacco odor. 

Melzer's test: red color, heated with 2-3 cc. of epichlorohy- 
drin. 

Schindelmeiser's test: nicotine, after standing several hours 
with a drop of formaldehyde solution, gives an intense red color 
with a drop of cone. HNO3. 

Roussin's test: ether solution of iodine after some time pro- 
duces ruby red, crystalline needles. 

Aniline. — Yellow, reddish or brownish oil drop^ from evapora- 
tion of ether extract. (See page 56. ** Synopsis of Group I" 
for further details.) 

* Cantharidin is taken up in Chapter IV of this book upon page 196. Ether 
extracts this compound from acid solution but it dissolves with diflScuIty in this 
solvent (o.ii : 100 at 18**). 



NON-VOLATILE POISONS 137 

Veratrine. — Cone. H2SO4: soluble with yellow color, gradually 
changing to orange, then to red and finally to cherry red. Gen- 
tle heat hastens these changes. Solution at first shows greenish 
yellow fluorescence. 

Froehde : same color changes as with cone. H2SO4. 

Cone. HCl: very stable red color when heated in test-tube 
upon water-bath. 

Weppen's test: mixed with 6 times the quantity of cane 
sugar + a few drops of cone. H2SO4, gradually becomes green 
and finally blue. Cone. H2SO4 containing furfurol may be 
used instead. 

Vitali's test: same as for atropine (see below). 

Strychnine. — Fine, crystalline needles having a very bitter 
taste upon evaporation of ether extract. 

Oxidation test: colorless solution in cone. H2SO4 becomes 
evanescent blue or blue-violet with a little solid K2Cr207. 
Same color given by Mandelin's reagent but more permanent. 

Brucine. — Cone. HNO3: dissolves with blood red color soon 
changing to reddish yellow and yellow. Dilution of yellow 
solution in a test-tube with a little water and addition drop by 
drop of dilute SnCU + Aq changes yellow to violet. 

Careful addition of solution in dil. HNO3 to cone. H2SO4 as 
upper layer produces red or yellowish red zone. 

Atropine. — Vitali's test: evaporated upon water-bath in por- 
celain dish with a little fuming HNOa, gives yellowish residue 
which becomes violet when moistened with alcoholic KOH. 
Hyoscy amine and scopolamine also give this test. Strychnine 
and veratrine behave similarly. 

Physiological test: enlargement of pupil of eye caused by a 
single drop of solution i : 130,000. 

Cocaine. — Free base, precipitated by KOH -f- Aq from not 
too dilute cocaine salt solution, forms oil drops soon becoming 
solid and crystalline. 

Benzoyl group: heat 5 minutes in a test-tube upon boiling 
water-bath with i ce. cone. H2SO4. Odor of methyl benzoate 
upon addition of 2 cc. of water. Upon cooling, benzoic acid 



138 DETECTION OF POISONS 

separates. This acid, washed and dried, recognized by melting 
point (120°) and by tendency to sublime. 

Physiological test: anesthesia of the tongue. 

Codeine. — Cone. H2SO4: soluble without color. Reddish or 
more bluish upon long standing, or at once upon gentle wanning. 

Oxidation: Deep blue or blue-violet, when warmed with 
cone. H2SO4 and KH2ASO4, or with a little FeCU+Aq. 

Froehde: yellowish color soon changing to green and to blue 
upon gentle warming. 

Sugar test: purple-red color upon gently warming with cone. 
H2SO4 and a little cane sugar. Due to furfurol formed. 

Formalin test: dissolves in cone. H2SO4 containing formalde- 
hyde with reddish violet color soon changing to permanent blue- 
violet. 

Pellagrins test: given by codeine (see apomorphine, page 140). 

Hydrastine. — Froehde: dissolves with fairly permanent green 
color later changing to brown. 

Mandelin: dissolves with reddish color gradually changing 
to orange-red. 

Fluorescence: intense blue fluorescence (characteristic) upon 
shaking dil. H2SO4 solution with very dilute KMn04 + Aq added 
carefully drop by drop. 

Quinine. — Amorphous varnish having very bitter taste from 
ether. 

Fluorescence: blue fluorescence in dil. H2SO4. 

Thalleioquin test: emerald green color, upon adding i cc. satu- 
rated Cl2 + Aq to dilute acetic acid solution and then at once 
excess of (H4N)0HH-Aq drop by drop. 

Herapathite test: heat to boiling with 10 drops of mixture 
(30 drops acetic acid + 20 drops absolute alcohol + i drop dil. 
H2SO4) and add i drop alcoholic iodine solution (1:10). 
Shining, olive green leaflets, appearing cantharides green by 
reflected light. 

Antipyrine. — Freely soluble in water. Neutral. Mildly 
bitter. 

Dissolve ether residue in little water and test for antipyrine as 
directed in A (see page 135). 



NON-VOLATILE POISONS 



139 



I 



Pyramidone.— Fine needles from ether. Freely soluble in 
Water. Neutral. 

FeClj + Aq: aqueous solution blue-violet or more red-violet. 

HNOj: f umi ng acid renders aqueous solution blue to blue- 
violet. 

Caffeme. — Concentric clusters of shining needles from ether. 
Mild, bitter taste. Fairly soluble in water. Neutral. 

Apply tests described under A (see page 136). 

Physostigmine.— (H,N)OH-i-Aq: evaporated with (H4N)- 
OH + Aq, gives blue residue soluble in alcohol with same color. 

Physiological test: causes contraction of pupil of eye. 

Narcotine. — Not bitter. Neutral. 

Froehde: soluble with green color. A concentrated reagent 
(0.05 gram (H4N)2Mo04 to i cc. cone. H^SOi) gives greenish 
color at first which gradually changes to cherry red and to a 
blue from margin toward center. 

Erdmann: soluble with fine red color. 

Papaverine.' — Tasteless, colorless, neutal prisms. 

Cone. HjSO*: pure alkaloid soluble without color. Heat 
produces dark violet color. 

Froehde: soluble with green color, soon changing when warmed 
to blue, violet and finally cherry red. 

HNOj-HsSO* test: cone. HjSO* containing HNOj, or cone. 
HNOj itself, gives a dark red solution. 

Thebaine. — Tasteless, colorless, alkaline prisms. 

Cone, H2SO1: soluble with deep red color. Froehde and 
Erdmann behave similarly. 



I 



C. Ether Extract' of Ammonia Solution may Contain: 

Apomorphine. — Residue amorphous and usually green. 
HjSO^-HNOj'test: solution in cone. HjSO^ colored evanescent 
violet, then reddish yellow or orange by drop of cone. HNOa. 
Froehde: soluble with green or violet color. 
Pellagri's test: dissolve in dil. HCl, add excess of NaHCOg, 



'Unless the tarlarii 
u described c 
BJt ttm extraction. 



add and alkaline solntions, as veil as tbar ether extracta, 
1 page tit, that is to say, have a green or red color, 



140 DETECTION OF POISONS 

shake well and add 2 drops alcoholic iodine solution. Blue or 
emerald-green color soluble in ether with violet color. 

Wangerin's test: 1-2 drops K2Cr207 + Aq (0.3 per cent.), 
added to apomorphine hydrochloride solution, gradually pro- 
duces dark green color. Chloroform added becomes violet. 
Addition of dil. SnCU + Aq produces pure indigo-blue color. 

D. Chloroform Extract of Ammonia Solution may Contain : 

Morphine. — Very bitter. Usually amorphous. Rarely crys- 
talline. 

Froehde: soluble with violet color gradually changing to 
dirty green and finally to pale red. 

Formaldehyde-H2S04: soluble with purple-red color later be- 
coming blue-violet and almost pure blue. 

Husemann's test: dissolve in cone. H2SO4, heat over very small 
flame until abundant white fumes appear, cool and add i drop 
cone. HNOs. Very evanescent, red-violet color which soon 
changes to blood-red or reddish yellow. 

Pellagrins test: see apomorphine. 

FeCls+Aq: dissolve in few drops very dilute HCL evaporate 
to dryness upon water-bath, dissolve in little water and add 
drop FeCla+Aq. Blue color. 

Bismuth test: dissolve in cone. H2SO4 and sprinkle bismuth 
subnitrate on surface of solution. Dark brown color. 

Antipyrine and Caffeine. — Being soluble in ether with some 
difiiculty, but readily soluble in chloroform, these substances 
may appear in the residue from D, if they have not been pre- 
viously completely extracted with ether. 

Narceine. — 12 test: blue color with I2 + Aq. 

Resorcinol-H2S04 test: dissolves in resorcinol-H2S04, giving 
intense yellow solution which becomes carmine-red or cherry- 
red, if warmed upon the water-bath and stirred. 

Tannin-H2S04 test: dissolves in tannin-H2S04, giving yellow- 
ish brown solution which becomes pure green* if warmed upon 
the water-bath. 



t 1 



CHAPTER III 
METALUC POISONS 

Destruction of Organic Matter 

The analyst cannot rely upon tests for poisonous metals, if 
animal or vegetable matter is present. Consequently complete 
destruction of interfering organic substances is absolutely 
essential to success. Description of a few of the more im- 
portant methods used for this purpose will suffice. 

I. Fresenius-v. Babo Method^ 

The residue left after removal of volatile poisons by steam 
distillation may be used in this part of the analysis, as it must 
contain poisonous metals if any are present. 

A portion of the original material,^ previously finely chopped 
and well mixed in a large flask with enough water to produce a 
fluid mass, may also be used. According to the quantity of 
material, add lo, 20 or 30 cc. of pure concentrated hydrochloric 
acid.* Finally add 1-2 grams of potassium chlorate, shake well 
and set the flask upon a boiling water-bath. Nascent chlorine 
should come into contact with the material as intimately as 
possible. 

When the mixture is hot enough, add 0.3-0.5 gram of potas- 
siiun chlorate at 5 minute intervals and shake the flask fre- 
quently. Continue in this manner, until most of the organic 
matter is dissolved and the solution is pale yellow. Further 

' Annalen der Chemie und Pharmazie 49, 306 (1844). 

^ Cadaveric material should be divided as finely as possible, then brought to a 
thin mixture by stirring with 12.5 per cent., arsenic-free hydrochloric add and 
heated with frequent shaking with 1-2 grams of potassium chlorate as directed 
above. If the material is heated on the water-bath in a porcelain dish, it should 
be stirred constantly. 

* In laboratory experiments 5-10 cc. cone, hydrochloric add is usually suffi- 
cient. A large excess of hydrochloric add should be avoided. 

141 



142 



DETECTION OF POISONS 



addition of potassium chlorate and longer heating should 
produce no real change. Fat especially resists the action of 
chlorine. 

When organic matter is completely destroyed, dilute with 
hot water, adding a few drops of dilute sulphuric add to pre- 

cipitate possible barium, 
shake and pour the liquid 
through a wetted filter. If 
the excess of free hydro- 
chloric acid is not too large, 
saturate the filtrate direct 
with hydrogen sulphide as 
directed on page 145 . Other- 
wise, evaporate the solution 
in a porcelain dish upon the 
water-bath nearly to dryness 
to remove most of the free 
hydrochloric add. This step 
frequently gives rise to a dark 
brown color which a few 
crystals of potassiimi chlorate 
will discharge. In testing for 
lead, cadmiimi and copper, it 
is advisable to evaporate, 
because hydrogen sulphide 
predpitates the first two 
metals incompletely, or not 
at all, from solutions contain- 
ing too much hydrochloric 
acid. 
An alternative procedure consists in removing part of the 
free hydrochloric add from the filtrate, obtained after treat- 
ment with hydrochloric acid and potassium chlorate, by first 
evaporating to smaller volume and then adding ammonium 
hydroxide solution until alkaline. Add dilute nitric acid until 
the solution is faintly add and saturate with hydrogen sulphide 
(see page 145). 




Fig. 12. 



METALLIC POISONS 



143 



I The residue upon the filter may contain silver chloride, 
barium sulphate and lead sulphate in addition to fat. Examine 
as directed under "Metallic Poisons IV" (see page 163}. 

H. Thorns' destroys organic matter in the apparatus shown 
in Fig. 12. Oxidation is carried on in an ordinary fractioning 
flask (A) with the tubulus (B) bent upward. A separating 

i funnel (C), held in the neck of the flask by a stopper, contains 
an aqueous solution of potassium chlorate (i : 30) saturated at 
room temperature, The organic matter is in the flask as a 
thin mixture with 12.5 per cent, hydrochloric acid. Add about 
1 gram of solid potassium chlorate and warm the flask on a 
boiling water-bath. When the mass in the flask is warm, let 
the potassium chlorate solution run in drop by drop and shake 
constantly. Care must be taken not to add too much of this 

PBolution at once; otherwise the procedure is identical with that 
|»eviously described. 
Notes. — Potassium chlorate and hydrocbloric add evolve chlorine (a and 0) • 
pait of which acts upon the orgaoic material and part in contact with water formi 
oxygeo and oxygen acids of chlorine (HOCl) [y and i) which are strong oxididng 

(«) KCIO, + HCl - HCIO, + KCl. 

{&) HCIO, + sHC! = 3CI, + 3H,0, 

(y) Cli + H,0 = jHCt + O, 

(i) Cl, + H,Oi=: HOCl + HCl. 

Vhlte ArMnlc (AiiOi) in a mixture probably cannot be volatilized us arsenic 
trichloride (.AsCliJ in the procedure described but is oxidized to non-volatile 
Arsenic add (HiAsOO: 

As,0, + iH,0 -t- sCI, = AsiO, + 4HCI. 

As,0, -I- 3H1O = iH.AsO,. 

There always remains, even after the most thorough treatment with hydro- 
chloric add and potassium chlorate, an insoluble white residue wholly unaffected 
by the action of chlorine. This is the case, espedally after the oxidation o( 
vegetable substances or cadaveric materia!. This treatment converts a pordon 
of the organic matter into volatile compounds (cbloranil?) which have a sharp 
odor and attack the mucous membranes. For this reason destruction of organic 
matter should take place in a hood with a good draft. 

Treatment as described with hydrochloric add and potassium chlorate converts 
metallic poisons into inorganic salts, usually chlorides and sulphates. These 
dtber remain in solution or appear as predpilates (AgCI and BaSOi). Protein 
substances, present in all animal and vegetable organisms, predpitate many 

' H. Thorns, "Einfiihrung in die praktischeNahrung5mittelChemie,"Ldpa8, 
i8g9. Published by S. Hir^el, Leipzig, iSqq. Abbtldung 64, page 153. 



144 DETECTION OF POISONS 

heavy metals, as mercury, silver, lead, copper and zinc from solutions of their 
salts. These metals are then in the form of metallic albuminates, some of which 
dissolve in water with great difficulty and are very stable. Usually these metal- 
lic protein compounds must receive further treatment before it is possible to 
detect the metal. Many organic adds, as tartaric add, and carbohydrates inter- 
fere more or less with the detection of heavy metab. In combination with these 
organic substances heavy metals are like copper in potassium cuprocyanide 
(K4Cus(CN)e), which ndther sodium hydroxide nor hydrogen sulphide will pre- 
cipitate because it is electrolytically dissociated in solution in part as follows: 

K4Cu,(CN)« ?=fe 4K- -f Cu,(CN)t"". 

In other words, the solution does not contain cuprous ions. If potassium cupro- 
cyanide is heated with hydrochloric add and potassium chlorate, copper passes 
into solution as cupric chloride. The reagents mentioned above now predpitate 
copper, for cupric chloride ionizes as follows: 

CuCl2?=±Cu- + 2CI'. 

and the solution now contains cupric ions. 

The detection, therefore, of these metallic poisons by the usual ionic reactions 
requires a prodecure which permits the analyst to bring about complete destruc- 
tion of interfering organic substances. The metals in question are thus converted 
into inorganic salts. 

Potassium chlorate acts best only in strong hydrochloric acid solution. Conse- 
quently this add should always be in excess. If the mass becomes too thick at 
any time during heating, it should be diluted \^dth water or dilute hydrochloric 
add. Also the contents of the flask should be well shaken during treatment with 
potassium chlorate, to prevent a large quantity of this salt from collecting upon 
the bottom of the flask. Such an occurrence may cause an explosion due to 
formation of the exceedingly unstable dioxide of chorine (ClOt).' 

The author employs in such analyses 12.5 per cent, hydrochloric add (sp. gr. 
1. 061), saturated with hydrogen sulphide and kept in a loosely stoppered bottle. 
This insures predpitation of the final traces of arsenic sometimes present even in 
the purest commerdal acid. Before being used, this add is filtered through ash- 
free paper to remove precipitated sulphur which may contain arsenic sulphide. 

Cadaveric material, heated i^ith hydrochloric add and potassium chlorate, is 
dissolved rather easily. An experiment, in which 100 grams of stomach and 
duodenum, 20 grams of stomach contents, 75 grams of kidney and 200 grams of 
liver (in all 395 grams) were treated as described, required about i hour for com- 
plete solution. The insoluble part was collected upon a filter and washed. It 
was amorphous, gummy, yellowish white and greasy. After being dried upon 
a porous earthen plate, it weighed 52 grams. Dned at 100®, it weighed 
only 32 grams. 

2. Sonnenschein-Jeserich Method 

This method requires the use of pure free chloric acid instead 
of potassium chlorate. Place the finely divided material in a 

» (a) KCIO, + HCl = HCIO, + KCI, 
(^) 3HCIO, = HCIO4 + 2CIO2 + H,0. 



METALLIC POISONS 145 

laxge flask and dilute with water. Add a few cc. of chloric acid 
and warm slowly and cautiously upon the water-bath. As soon 
as the mass swells and becomes porous, gradually add small 
portions of hydrochloric acid. Even a considerable quantity 
of cadaveric material will dissolve in 2-3 hours. Water lost 
by evaporation should be replaced occasionally, otherwise the 
reaction may take place with explosive violence. In other re- 
spects, the product of the reaction should be treated as already 
described. 

3. C. Mai's Method ^ 

Mix the finely divided material with dilute hydrochloric acid 
(i : 12) until thin. Add a little potassium chlorate and heat 
over a free flame, adding from time to time small quantities of 
potassium chlorate (0.2 gram). Cool as soon as liquefaction of 
the mass is complete. Fat separates and usually can be re- 
moved easily from the liquid. Heat this fat once or twice with 
very dilute nitric acid, filter and add the filtrate to the main part 
of the liquid. Continue heating the latter, adding small quan- 
tities of ammonium persulphate, (H4N)2S208, until the liquid is 
clear and light yellow. Filter and saturate the filtrate as usual 
with hydrogen sulphide. Ammonium persulphate is a power- 
ful oxidizing agent and also adds nothing non-volatile to the 
liquid. 

Examination of Filtrate for Metallic Poisons 

Predpitfition by Hydrogen Sulphide 

A solution properly prepared according to the Fresenius-v. 
Babo or any other method, freed from excess of hydrochloric 
acid and filtered, should have only a faint yellow color. ^ Heat 
such a solution in a flask upon the water-bath and saturate 
with arsenic-free hydrogen sulphide.^ Pass hydrogen sulphide 

^ Zdtschrif t iili Untertersuchung der Nahrungs- und Genussmittel 5, 1106 

(1902). 

'Chromium in not too small quantity imparts more or less of a green 
color both to the solution and the filtrate from the hydrogen sulphide precipitate, 
owing to the presence of chromic chloride (CrCU). 

' Prepare arsenic-free hydrogen sulphide by saturating dilute sodium hydroxide 
solution with hydrogen sulphide from crude iron sulphide and commercial hydro- 
10 



146 



DETECTION OF POISONS 



for 0,5-1 hour or longer' into the hot solution and continue 
this treatment after the solution has been removed from the 
water-bath and is cold. 

Allow the solution saturated with hydrogen sulphide to stand 
in the loosely stoppered flask for several hours or until the next 
day. If the solution then smells of hydrogen sulphide and 
blackens a piece of lead acetate paper held over it, the next stqi 




Fig. 13.— Apparatus for Generaticg Arsenic-free Hydrogen Sulphide, (a) 
Generator with dilute sulphuric add; <£) Separating funnel with NaSH; (c) Wash- 
bottlei (d) Solution to be saturated with HiS. 



in the process may be taken. Otherwise warm the solution 
once more upon the water-bath and again saturate with hydro- 
gen sulphide. Finally collect the hydrogen sulphide precipi- 
tate upon a small paper and wash with hydrogen sulphide water. 
Examine the precipitate for arsenic, antimony, tin, mercury, 
lead, copper, bismuth and cadmium (Metallic Poisons I and II) 

chloric add. Pour this sodium hydro-sulphide (NaSH) solution into a separating 
funnd and add slowly to dilute sulphuric add (1:4). The generation of the gas 
can be carried on in the apparatus shown in Fig. 13. 

' In laboratory experiments treatment with hydrogen sulphide may be shortened 
somewhat. A Kipp generator in which the gas is prepared from iron sulphide 
and hydrochloric add may be used. 



METALLIC POISONS 



147 



' EUid the filtrate from this precipitate for chromium and zinc 
(Metallic Poisons III). 

Vegetable and animal substances, after treatment with 
hydrochloric acid and potassium chlorate, frequently give 
liquids yielding colored precipitates' with hydrogen sulphide 
even in the absence of the metals mentioned above. Such 
precipitates consist largely of organic sulphur compounds. 
Consequently, if hydrogen sulphide produces such a colored 
precipitate in acid solution, it is not final proof of the presence 
of a metallic poison. Also without further examination it is 
impossible to decide from the color of the hydrogen sulphide 
precipitate as to the presence of a particular metal. 

Complete Precipitation. — Before testing for chromium and 
zinc in the filtrate from the hydrogen sulphide precipitate, add 
about lo times the volume of strong hydrogen sulphide water 
to a small portion of the solution, stir well and let stand several 
minutes. Unless a colored precipitate appears, the metals in 
question (Metallic Poisons I and II) have been completely 
removed and the filtrate may then be further tested for chrom- 
ium and zinc (Metallic Poisons III), Otherwise, first dilute 
the entire filtrate from the hydrogen sulphide precipitate with 
water and again saturate with hydrogen sulphide. In presence 
of much hydrochloric acid, lead and cadmium are incompletely 
precipitated by hydrogen sulphide (sec above). 

Treatment of Hydrogen Sulphide Precipitate with Ammonia 
and Yellow Ammoniimi Sulphide. — Extract the thoroughly 
washed hydrogen sulphide precipitate while still moist upon the 
filter with a hot mixture of appro.ximately equal parts of am- 
monia and yellow ammonium sulphide. Heat about 5-10 cc. 
of the mixture of ammonia and yellow ammonium sulphide to 
boiling and drop the solution over the precipitate upon the filter. 
Reheat the filtrate and again pour over the precipitate. Repeat 
this operation several times. Finally wash the filter with a 
' Repeated treatment with potassium chlarale and hydrochloric acid dissolves 
thoroughly washed casein and Shrin almost completely and gives a filtrate (lom 
which hydrogen sulphide precipitates dirty yellow to hrownlsh substances. 
These products are amorphous and contain organic sulphur compounds together 
piritb much free sulphur. 



148 DETECTION OF POISONS 

few cc. of a fresh mixture of ammonia and yellow ammonium 
sulphide. Test the entire filtrate for arsenic, antimony, tin 
and copper^ = Metallic Poisons I. Test the residue upon the 
filter for mercury, lead, copper, bismuth and cadmium = Me- 
tallic Poisons II. 

METALLIC POISONS I 

Examination of the Part of the Hydrogen Sulphide Precipitate Sdioble 

in Axmnoma-Ammonium Sulphide 

Arsenic, Antimony, Tin, Copper 

Use the solution prepared as described by treating the hydro- 
gen sulphide precipitate with a hot mixture of ammonia and 
yellow ammonium sulphide. This solution is usually dark 
brown owing to dissolved organic substances.* Evaporate the 
solution to dryness in a porcelain dish upon the water-bath. 
Moisten the cold residue with fuming nitric add and again 
evaporate. Then intimately mix the residue with about 3 times 
its volume' of a mixture of 2 parts of sodium nitrate and i part 
of dry sodium carbonate. Thoroughly dry this mixture upon 
the water-bath and introduce small portions at a time into a 
porcelain crucible containing a little fused sodium nitrate 
heated to redness. After the final addition, heat the crucible 

^ Copper sulphide (CuS) is somewhat soluble in hot yellow ammonium sul- 
phide. An ammonium sulphide solution containing copper, treated as described 
on page 149, yields copper oxide which gives the melt a more or less gray or 
black appearance. If the melt is extracted with water, the residue contains 
black copper oxide with stannous oxide and sodium pyro-antimonate. To detect 
copper, dissolve the black residue in a little hot dilute hydrochloric acid and divide 
the solution into two parts. Add ammonia to i part until alkaline. The solu- 
tion is blue, if copper is present. Add potassium ferrocyanide solution to the 
other part. A brownish red precipitate of cupric ferrocyanide (CufFe(CN)o) 
appears, if copper is present. 

* In absence of metals the appearance of dark colored precipitates (see above), 
when the hydrochloric acid solution is treated with hydrogen sulphide, should not 
be misunderstood. Such precipitates are due to organic substances soluble with 
a dark brown color in a hot mixture of ammonia and yellow ammonium sulphide. 

* In most laboratory experiments 3 grams of a mixture of 2 grams of sodium 
nitrate and i gram of sodium carbonate is sufficient. A large excess of sodium 
nitrate should be avoided. 



METALLIC POISONS 149 

a short time, introducing possibly a little more sodium nitrate, 
until the fused mass is colorless. In presence of copper the 
melt is gray or grayish black from copper oxide. Sodium 
arsenate, sodium pyro-antimonate, sodium stannate, as well as 
stannic oxide and copper oxide, may also be present. Soften 
the cold melt with hot water and wash into a flask. Add a 
little acid sodium carbonate to the clear or cloudy liquid to de- 
compose the small quantity of sodium stannate possibly in solu- 
tion and precipitate all the tin as stannic oxide and then filter. 
The filtrate (A) contains any arsenic present as sodium arsen- 
ate (Na2HAs04) and the residue upon the filter (B) may con- 
tain^ sodium pyro-antimonate (Na2H2Sb207) , stannic and copper 
oxides. 

Examination of Filtrate A for Arsenic 

Arsenic is isolated as the element. Positive proof of the pres- 
ence of the poison is thus afforded. Two methods are in use for 
this purpose, namely, the Marsh-Berzelius and the Fresenius- 
V. Babo method. Both are very accurate and exclude any 
confusion of arsenic and antimony. 

Chapter V gives the details for detecting arsenic by the very 
delicate biological test, which requires the use of certain moulds, 
and also for the elctrolytic separation of arsenic at the cathode 
as arsine. 

I. Marsh-Berzelius Method 

Principle. — Nascent hydrogen converts oxygen compounds 
of arsenic, arsenious and arsenic acids, as well as arsenites and 
arsenates into arsine, AsHg: 

As20i 4- 12H = 2AsHi + 3H2O, 

AsjOft-aHjO* 4- 16H = 2AsH, -f- 8H2O. 

At a red heat arsine is decomposed into metallic arsenic and 

hydrogen : 

AsH, = As + 3H. 

This reaction represents the formation of the arsenic mirror. 

^ Even in the absence of the substances mentioned under B, a small insoluble 
residue usually appears. This may come from the porcelain crucible, the glazing 
of which is slightly attacked in the fusion with sodium nitrate and carbonate. 

' AstO».3HiO is the dualistic method of writing 2H1ASO4. 



150 DETECTION OF POISONS 

Also hydrogen containing arsine burns with a bluish white 
flame (a) . Depress a piece of cold porcelain upon such a flame. 
Hydrogen will burn but a deposition of metallic arsenic takes 
place (/3). This is the so-called arsenic spot: 

(a) 2AsH, + 3O1 = As,0» + 3H1O, 
05) 2AsHt + 3O « 2As + 3H1O. 

Hydrogen containing arsine precipitates black metallic silver, 
if passed into dilute silver nitrate solution. The solution con- 
tains arsenious add: 

AsH, + 3H2O + 6AgN0, = HjAsO, + 6HN0, + 6Ag. 

Procedure. — First acidify " Filtrate A," prepared as described 
(page 149) and possibly containing sodiiun arsenate, with dilute 
arsenic-free sulphuric add. Evaporate this solution in a porce- 
lain dish upon an asbestos plate over a small free flame. Add a 
few drops of concentrated sulphuric add to expel completely 
any nitric add possibly present in the residue and heat until 
copious white fumes of sulphuric add appear. The residue* 
in the porcelain dish is a thick colorless liquid having a strong 
add reajction. Arsenic, if present, is in the form of arsenic add 
which when cold frequently solidifies to a white crystalline 
mass. Examine the solution of this residue in the Marsh ap- 
paratus for arsenic. The same solution may also be used in 
testing for arsenic electrolytically (see pages 226 and 231). 

Marsh Apparatus. — Place 30-40 grams of pure arsenic-free 
zinc^ (granulated or in small rods) in the reduction flask A of 
the Marsh apparatus (Fig. 14). Pour cold dilute arsenic-free 
sulphuric add upon the metal. This add should contain 
15-16 per cent, of H2SO4.' Control the temperature of the 
solution, which should not rise much during the analsrsis, by 

^ To insure complete removal of nitric add, test a few drops of this residue with 
ferrous sulphate and sulphuric add. 

*The passage of hydrogen from 15-20 grams of zinc, treated with dilute 
arsenic-free sulphuric add, through the strongly heated ignition tube C of the 
Marsh apparatus should not give a trace of arsenic after i hour. The metal is 
then pure enough for use. Bertha spdter from the New Jersey Zinc Company 
will meet such a test. 

* Add I volume of pure arsenic-free concentrated sulphuric add to 5 volumes 0/ 
distilled water. This diluted add when cold is suitable for use in the Marsh test. 



METALLIC POISONS 



151 



' generating hydrogen slowly. Otherwise, there is danger of 
partial reduction of sulphuric add to sulphur dioxide and then 
to hydrogen sulphide which interferes more or less with the 
detection of arsenic. Place the reduction flask A in a dish of 
cold water, if the add becomes too warm. 

Certain precautions are necessary in using the Marsh ap- 

Iiratus. 
1. Have the apparatus absolutely tight. 
2. Expd air completely before igniting hydrogen. To tell 
hen this point is reached, collect hydrogen in a dry test-tube 




[ Fig. 14' — Marsh Appiiratua. (a) Hydrogen-generator; (A) Chloride of caI- 
n drying-tube; (e) Hard glass tube; id) Arsenic it 



until it ignites without detonation when carried to a flame. If 
the hydrogen stands this test, ignite the gas at the end of igni- 
tion tube C. There is no danger of an explosion within the 
apparatus. If the apparatus is tight and the evolution of 
hydrogen is not too rapid, it requires about 8 minutes to expd 
_ the air. 

3. Test the hydrogen to insure its entire freedom from arsenic, 
^either the arsenic mirror nor spot appears. 
I If the hydrogen is arsenic-free, gradually introduce the 
rfectly cold sulphuric add solution, containing arsenic as 
rsenic add (page 150), in small portions into reduction flask A, 
j^t the same time heat ignition tube C to redness just back of 



152 DETECTION OF POISONS 

the capillary tube. If the solution contains arsenic, the gas 
generated consists of a mixture of arsine (AsHs) and hydrogen. 
A shining mirror of metallic arsenic appears, often in a few 
minutes, just beyond the point of ignition. Traces of arsenic 
require considerable time before a brown or brownish black 
j&lm appears. A piece of white paper held behind the tube 
brings out clearly even a minute arsenic mirror. 

Remove the flame from ignition tube C. If arsenic is present, 
the hydrogen flame becomes bluish white. At the same time 
white fumes of arsenious oxide (AS2O8) arise from the flame. 

To produce the lustrous, brownish black spot (arsenic spot), 
depress a cold porcelain dish upon the hydrogen flame. 

Arsine has an exceedingly characteristic, garlic-like odor. 
Extinguish the hydrogen flame and allow the gas to escape.^ 
The odor is evident even when the hydrogen contains traces 
of arsine. 

A third method of detecting arsenic by the Marsh apparatus 
consists in extinguishing the hydrogen flame and passing the 
gas into dilute silver nitrate solution. Arsine darkens this 
solution, producing a black precipitate of metallic silver. The 
solution contains arsenious acid and free nitric acid. (See 
reaction, page 150.) 

Filter through a double paper to remove silver and carefully 
neutralize the filtrate with a few drops of very dilute ammonium 
hydroxide solution. If the solution is neutral, it is possible 
to obtain a yellowish white precipitate of silver arsenite 
(AgsAsOa) but this compound dissolves easily in ammonium 
hydroxide solution and in nitric acid. 

Extinguish the flame at the end of reduction tube C and hold 
over the tube a strip of paper moistened with concentrated 
silver nitrate solution (1:1). A yellow stain appears, if the 
hydrogen contains arsine. A drop of water added to this 
yellow spot changes the color to black. This is Gutzeit's 
arsenic test (see page 156). 

' The detection of arsine by other tests is so easy that it seems somewhat 
superfluous to confirm its presence in this way in view of its very poisonous 
properties. Tr. 



METALLIC POISONS 



I 

1 

t 



DifEerences Between Arsenic and Antimony Spots and Miirors 

Nascent hydrogen reduces various antimony compounds (SbGi, SbtOi, 
HSbO., KSbOCiHjO,, etc.) producing the colorless gas sUbine (SbHi). Tlic 
behavior of this compound in the Klarsh apparatus closely resembles that ol 
arsine. tor it gives a spot and minor, and precipitates black silver antimonide 
(AgiSb) but not metallic silver, if passed into silver nitrate solution. 

The procedure employed in preparing materia! for the Marsh test (see page 
148) separates arsenic from antimony and eicludea the possibility of the two met- 
als appearing in the Marsh test at the same time. Since the identiiicalioD of 
arsenic mirrors and spots by other tests is important, the differences between ar- 
senic and antimony should be pointed out. The suspicion that antimony is 
present often necessitates other confirmatory tests (see Antimony, page 157). 
Introduce the solution into the Marsh apparatus and produce the antimony 
spot and mirror. 

The differences between arsenic and antimony spots and mirrors are: 

I. The arsenic mirror has a high metallic luster. It is brownish black and 
volatile. Owing to this latter property, it sublimes easily when heated in a 
stream of hydrogen. In the case of the antimony mirror, which appears on both 
sides of the flame, the metal in contact with the heated glass fuses and is silver 
white. But in those places removed from the flame it is almost black and has 
hardly any luster. Slibine decomposes at a temperature much below that 
re<]uired for arsine. This fact explains the deposition of liiis metal on both sides 
of the Same. Antimony volatilizes at a high temperature and consequently 
■ublimea with difficulty. 

The arsenic spot, if not too heavy, is brownish black or brown and lustrous. 
^Jt dissolves readily in sodium hypochlorite solution, forming arsenious add; 
3H,0 + jAs + jNaOCl - 2H,.\sO, + jNaCl. 

The antimony spot is dull, velvet black and without luster. A thin film of 
antimony is never btown but has a dark, graphite-like appeajance. It is insolu- 
ble in sodium hypochlorite solution, 

3. A drop of concentrated nitric add, or moist chlorine, at once dissolves the 
arsenic spot forming arsenic add. Neutralize with ammonia and add silver 
nitrate solution. A reddish predpitate of silver arsenate (AgiAsOi) appears. 

Nitric add, or moist chlorine, also dissolves the antimony spot but silver 
nitrate does not produce a colored predpitate. 

4. Gently heat the ignition tube and pass a stream of dry hydrogen sulphide 
over the arsenic mirror. Yellow arsenic trisulphide (AsjSi) appears. The 
antimony mirror becomes brownish red lo black (SbiSi). 

;. Ar^e passed into silver nitrate solution precipitates black metallic silver 
Mid the filtrate from such a predpitate contains aisenious add. But stibine 
predpitates black silver antimonide (AgjSb) and the filtrate does not contain a 
trace of antimony since the predpitation of black AgiSb is complete. To detect 
antimony, collect the black predpitate upon paper, wash and heat for some time 
in 10-15 per cent, tartaric acid solution. Antimony dbaolves, whereas sOver 
remaiiu as a grayish white residue. Add dilute hydrochloric add to this solu- 
tion and then treat with hydrogen sulptiide. Antimony appears as orange-red 
Antimony trisulphide. 



154 



DETECTION OF POISONS 



2. Fresenius-von Babo Metiiod 

Principle. — Fusion of oxygen and sulphur compounds of 
axsenic with a mixture of sodium carbonate and potassium 
cyanide causes reduction with formation of an arsenic mirror. 
As a result potassium cyanide changes to potassium cyanate 
(KCNO) or potassium sulphocyanate (KSCN) : 

As,0, + 3KCN = As, + aKCNO, 
AsiS, + 3KCN = As, + 3KSCN. 

Procedure. — Use for this test the sulphuric add solution 
prepared by the method already described and containing ar- 
senic in the form of arsenic acid (page 1 50) . To reduce arsenic 
add to arsenious add, add a few cc. of sulphurous add to the 
solution and heat until the odor of this add has disappeared. 







l>i*u;c :hi:i^ 5\>hxxvM\ wi;h wAttr A:>a :rx?A: with hydrogen 
l>huU\ vV.tUv: ?ho ivrtvVp^iu:^ v>: ironic :risci>cide 



r* X 



As,^^ 



£^ '=>» the 



VX vX^' 



I 
I 



METALLIC POISONS 155 

with a little water and add enough dry sodium carbonate to 
render the mixture distinctly alkaline. Dry thoroughly upon 
the water-bath and triturate the residue in a mortar with several 
times the quantity of a mixture of j parts of dry sodium car- 
bonate and I part of pure potassium cyanide. 

Transfer this mixture to a porcelain boat and place in an igni- 
tion tube of hard glass. Heat in a stream of carbon dioxide 
(Fig. 15) dried by means of arsenic-free sulphuric acid. 

To expel moisture, first heat the ignition tube gently where 
the boat is and then ignite at a bright red heat, A mirror 
of arsenic appears upon the cooler part of the tube, if arsenic is 
present. 

A simpler method of detecting arsenic by means of potassium 
cyanide is often used. Heat the thoroughly dried material 




I 



B, 

—(A) Substance and fusion-rai.Uure; (S) Arsenic ; 

containing arsenic (AsjOs, AsjSa) in a bulb tube with a dry mix- 
ture of sodium carbonate and potassium cyanide until fusion 
takes place. If the tube is smaller above the bulb, the arsenic 
mirror will form in the constricted area (Fig. 16}. 
Other Arsenic Teats 
1. Bettendorff's Test. — Concentrated stannous chloride solu- 
tion precipitates metallic arsenic from arsenious acid cold and 
from arsenic acid with heat or after long standing. This test 
requires the use of a special stannous chloride solution.' The 
solution is red to brownish red, if only traces of arsenic are pres- 
ent. More thaJi traces of arsenic produce a black precipitate of 
arsenic: 

As,0, + 6fICI -I- sSnCl, = i.\3 + 3H,0 + aSnCU. 

AsjO, -i- loHCl + sSnCU + jH^O = jAs + 8H,0 -|- sSnCI,. 

Use for this test the sulphuric acid solution obtained as de- 
scribed above (page 150) which contains arsenic in the form of 

1 See page 315 for the prcparalion of this reagent. 



156 DETECTION OF POISONS 

arsenic acid. Bettendorff's test is not as delicate as the Marsh 
test. 

2. Gutzeit's Test^ — This test permits the detection of minute 
traces of arsenious and arsenic acids, as well as their salts, with 
certainty. Generate hydrogen in a test-tube from arsenic-free 
zinc and pure dilute hydrochloric add. To remove any sul- 
phurous acid or hydrogen sulphide, add a few drops of iodine 
solution until the liquid is yellow. Then add the solution to be 
tested and place a loose cotton plug in the neck of the test-tube. 
If arsenic is present, the silver nitrate spot becomes lemon- 
yellow. 

AsHa + 6AgN0, = (sAgNCAgsAs) + 3HNO,. 

Gradually a brownish black border forms around the yellow 
spot. A drop of water at once turns the spot black from sepa- 
ration of metallic silver. 

(3AgN03.Ag,As) + 3H,0 = 6Ag + H,AsO, + 3HNO,. 

This is a very delicate arsenic test. One drop of o.i per cent, 
potassium arsenite solution produces a distinct yellow color 
upon the silver paper. Gutzeit's test is positive with even 0.05 
milligram of As203.^ The sulphuric acid solution containing 
arsenic as arsenic acid (see page 150) may be used for this test. 

But Gutzeit^s test is not as characteristic of arsenic as the 
Marsh test. Stibine, phosphine from phosphorus in zinc and 
even hydrocarbons color the silver nitrate paper. Dry hydro- 
gen sulphide also produces a yellow or yellowish green spot upon 
paper moistened with concentrated silver nitrate solution. 
The latter spot has a black border which gradually extends 
until the entire spot becomes black. Poleck gives this yellow 
compound the composition (AgNOs.AgaS). 

Detection of Antimony, Tin and Copper in Residue B 

Residue B (see page 149), insoluble in water and obtained 
from the fusion, may contain sodium pyro-antimonate, stannic 

* Gutzeit, Pharmazeutische Zeitung 1879, 263; and Poleck and Ttimmel, 
Berichte der Deutschen chemischen Gesellschaft 16, 2435 (1883). 

' See page 233 for the application of this method to the quantitative estimation 
of arsenic. Tr. 



METALLIC POISONS 157 

and cupric oxides. Treat this residue upon the filter with a 
little hot dilute hydrochloric acid (equal parts of concentrated 
add and water). Pass this acid repeatedly through the paper 
until most of the residue is dissolved. If the original color 
of residue B and of the melt was gray or black, first examine a 
portion of the hydrochloric acid solution for copper. Excess 
of ammonia produces a blue color. Potassium ferrocyanide 
solution gives a brownish red precipitate, or only a coloration 
with traces of copper. 

Concentrate the remainder of the hydrochloric acid solution 
to a few drops in a porcelain dish upon the water-bath; and put 
2 drops of this solution upon platinum foil in contact with zinc. 
Antimony produces a black, tin a grayish and copper a dark 
reddish brown spot upon platinum. There is little chance of 
confusing the tin or copper spot with that given by antimony. 

Dilute the remainder of the hydrochloric acid solution with 
water and introduce a piece of zinc. Keep the zinc in the solu- 
tion, as long as hydrogen is evolved. Collect the black, metal- 
lic flocks from this operation upon a small filter. Wash thor- 
oughly, and gently warm with a little concentrated hydrochloric 
acid. Finally, filter the solution. Antimony does not dissolve, 
whereas tin passes into solution as stannous chloride (SnCU), 
and is in the filtrate. Apply the tests described under tin to 
this solution. 



TIN 



The solution, treated as described in the above analytical 
procedure, contains tin as stannous chloride (SnCU). Apply 
the following tests for this metal: 

(a) Mercury Test. — Add to a portion of the filtrate a few 
drops of mercuric chloride solution. Tin precipitates white 
mercurous chloride (calomel). Heat produces in addition 
gray, metallic mercury, if there is a large excess of stannous 
chloride. 

(J) Prussian Blue Test. — Add to a second portion of the 
filtrate a few drops of a dilute mixture of ferric chloride and 



158 DETECTION OF POISONS 

potassium ferricyanide solutions. Tin produces a precii»tate 
of Prussian blue. 

This test b not characteristic of tin, as many other substances capable of 
reducing ferri-fenicyanide to fem-ferroc>'anide, that b to say, to Prussian bhie 
act in the same way. 

To identify antimony furtiier, dissolve tiie black flocks, in- 
soluble in hydrochloric add, in a few drops of hot aqua regia. 
Expel excess of add upon the water-bath and dilute the residue 
with water. If the quantity of antimony is not too small, water 
predpitates white antimony oxychloride (SbOCl). Redissolve 
this predpitate in a littie dilute hydrochloric add. Test a 
portion of this solution for antimony with hydrogen sulphide. 
Introduce the remainder into the Marsh apparatus and produce 
the antimony spot and mirror, or test for stibine with silver 
nitrate solution as described (see page 153). 

METALUC POISONS n 

Detection of Metals Whose Sulphides are Insoluble in Amm o niu m Sul- 
phide 
Mercury Bismuth 

Lead Copper 

Cadmium 

That portion of the hydrogen sulphide predpitate, insoluble 
in ammonium sulphide solution, may contain mercury, lead, 
bismuth, copper and cadmium sulphides. Examine this pre- 
dpitate according to the methods employed in qualitative 
analysis. Treat a small precipitate repeatedly upon the filter 
with a few cc. of warm, rather dilute nitric add (i volume of 
concentrated acid and 2 volumes of water). Mercuric sulphide 
does not dissolve, but the other sulphides pass into solution as 
nitrates. 

Detection of Mercury in the Residue Insoluble in Nitric Add 

Always examine that portion of the hydrogen sulphide pre- 
dpitate, insoluble in nitric add, for mercury, even when not 
black! Treat this residue upon the filter with a littie hot, 



METALLIC POISONS 



159 



I somewhat diluted hydrochloric acid, containing in solution a 
I few crystals of potassium chlorate and pass the acid through the 
I- paper several times. Evaporate the filtrate to dryness in a 
I porcelain dish upon the water-bath, and dissolve the residue in 
I a-3 cc. of water containing hydrochloric acid. Filter this solu- 
[ tion, and examine the filtrate for mercury. 

(a) Stannous Chloride Test.— Add to a portion of the filtrate 
I a few drops of stannous chloride solution. A white precipitate 
I of mercurous chloride (calomel) appears, if mercury is present, 
I Excess of stannous chloride, especially if heat is applied, re- 
I duces this precipitate to gray, metallic mercury. 

{b) Copper Test — Put a few drops of the filtrate upon a 
small piece of bright copper. Mercury immediately deposits 
a gray spot which has a silvery luster when rubbed. Wash 
I the copper, upon which mercury has been deposited, successively 
Lin water, alcohol and ether. Dry thoroughly and heat in a 
1 small bulb-tube of hard glass. Mercury sublimes and collects 
I in small, metallic globules on the cool sides of the tube. A 
I trace of iodine vapor, introduced into the tube immediately trans- 
I forms the gray sublimate into scarlet mercuric iodide (Hglj). 

(c) Phosphorous Acid Test. — Add to another portion of the 
I filtrate some phosphorous acid and warm gently. A white 
I precipitate of mercurous chloride (calomel) appears, if mercury 
mb present: 

iHgCl, + H,0 + H,PO, = Hg,Cl, + 2HCI + HjPOi. 

(d) Precipitation of Mercuriclodide. — Add 1-2 drops of very 
dilute potassium iodide solution to the remainder of the filtrate, 
A red precipitate (Hglj), readily soluble in excess of potassium 

^Liodide, shows mercury: 



ination of the Nitric Acid Solution 



The nitric acid solution may contain lead, bismuth, copper 
and cadmium nitrates. Evaporate this solution in a porcelain 
dish nearly to dryness and dissolve the residue in a little hot 



160 DETECTION OF POISONS 

water. If the solution contains lead, dilute sulphuric acid pro- 
duces a heavy white precipitate of lead sulphate. Test the 
filtrate from this precipitate for bismuth, copper and cadmium. 

(a) Copper and Bismuth Tests. — Excess of ammonium hy- 
droxide solution, added to most of this filtrate, produces a blue 
color if copper is present. A white precipitate at the same time 
may be bismuthous hydroxide^ (Bi(0H)8). To detect bismuth, 
wash the precipitate and dissolve upon the filter in a few drops 
of hot dilute hydrochloric acid. Pour this solution into consid- 
erable water. A white precipitate of bismuthous oxychloride 
(BiOCl) proves the presence of bismuth. As an alternative 
test, add stannous chloride to the hydrochloric acid solution and 
then excess of sodium hydroxide solution. A black precipitate 
of metallic bismuth appears. 

(b) Potassium Ferrocyanide Test. — Potassium ferrocyanide 
solution precipitates copper as brownish red cupric ferrocyanide 
(Cu2Fe(CN)6). Traces of copper produce a brownish red color. 
There is a deposit of cupric ferrocyanide after some time. 

(c) Precipitation of Metallic Copper. — A bright knife blade, 
or a bright iron nail, immersed for a short time in a copper solu- 
tion, becomes red from a coating of metallic copper. 

To detect cadmium in presence of copper, add solid potas- 
sium cyanide to the blue solution, produced by ammonium 
hydroxide, until the blue color is discharged. Then pass hydro- 
gen sulphide through the solution. Cadmium is precipitated as 
the yellow sulphide (CdS), whereas copper remains in solution 
as K4Cu2(CN)6, potassium cuprocyanide. 

When copper is absent, test for cadmium by passing hydrogen 
sulphide at once into the ammoniacal solution. If a reddish 
or brownish instead of a yellow precipitate appears, filter, dry 
the precipitate upon the paper and heat upon charcoal in the 
blow-pipe flame. Cadmium gives a brown coating. 

^ Ammonium hydroxide solution does not precipitate pure bismuthous hydrox- 
ide (Bi(OH)t) from solutions of bismuth salts but a basic salt, the composition 
of which depends upon the temperature and concentration of the particular 
solutions. 



METALLIC POISONS 161 

METALLIC POISONS m 
Detection of Chromiuni and Zinc 

Test the filtrate from the hydrogen sulphide precipitate for 
chromium and zinc. Concentrate the filtrate to about one- 
third its original volume and divide this solution into two parts. 

Detection of Zinc 

Add enough ammonium hydroxide solution to render one- 
half the concentrated filtrate alkaline. This treatment usually 
gives the solution a dark color. Then add ammonium sulphide 
solution in excess. This reagent almost always produces a 
precipitate even when zinc is not present, since solutions, pre- 
pared from animal and vegetable materials, usually contain 
iron compounds and phosphates of the metals of the earths and 
of the alkaline earths. When this precipitate has settled, add 
acetic acid until the solution has a faint acid reaction. Stir the 
mixture thoroughly, and allow it to stand for some time. The 
color of the precipitate becomes lighter, because acetic acid dis- 
solves sulphide of iron. Moreover, the phosphates are partly 
dissolved, except ferric phosphate (FeP04) which is insoluble in 
acetic acid. Collect the precipitate upon a filter. Wash, dry 
and ignite precipitate and filter in a porcelain crucible. Before 
ignition, moisten the filter with concentrated ammonium ni- 
trate solution. .Extract the residue from ignition with 3 cc. 
of boiling, dilute sulphuric acid. Filter, and divide the filtrate 
into two parts. 

(a) Add sodium hydroxide solution in excess to one portion 
of the filtrate and shake thoroughly. Filter, to remove the 
white precipitate of ferric phosphate which usually appears, and 
add a few drops of ammonium or hydrogen sulphide solution to 
the clear filtrate and heat. Zinc, if present, gives a white, 
flocculent precipitate of zinc sulphide. 

(b) Add ammonium hydroxide solution in considerable ex- 
cess to the second part of the filtrate. Filter, to remove ferric 

phosphate, and acidify the filtrate with acetic acid. Warm 
11 



162 DETECTION OF POISONS 

the solution and treat with hydrogen sulphide. Zinc, if present, 
appears as^the white sulphide. 

(c) Test]^further for zinc by dissolving the precipitate pro- 
duced by ammonium or hydrogen sulphide (as described above 
in a and ft), after it has been collected upon a filter and thor- 
oughly washed, in a few drops of hot dilute hydrochloric add. 
Boil until hydrogen sulphide is expelled, and filter to remove 
precipitated sulphur. Add potassium ferrocyanide solution'^to 
the clear, cold filtrate. This precipitates Zn2Fe(CN)e, zinc 
ferrocyanide, which is white, slimy and nearly insoluble in dilute 
hydrochloric acid.^ 

Detection of Chromium 

To test for chromium^ in the second part of the filtrate 
from the hydrogen sulphide precipitate, concentrate the solu- 
tion in a porcelain dish to a small volume. Then add twice the 
quantity of potassium nitrate and sodium carbonate, until the 
reaction is decidedly alkaline. Finally, heat this mixture until 
perfectly dry. Add this dry residue in small portions to a little 
potassium nitrate fused in a crucible. In fusing a large quan- 
tity of material, it is advisable to use a large, bright, nickel 
crucible which is especially adapted for this operation. When 
fusion is complete, cool thoroughly, boil crucible and contents 
with water in a porcelain dish and filter the solution. Chro- 
mium colors the filtrate more or less yellow. Even mere traces' 
of chromium color the filtrate yellow. When the solution of 
the melt is colorless, it is unnecessary to test for chromium. To 
detect chromium when the filtrate is yellow, divide the solution 
into two portions and make the following tests: 

^ Excess of potassium ferrocyanide combines with zinc ferrocyanide, which is 
first precipitated, and forms insoluble potassium zinc ferrocyanide: 

3Zn,Fe(CN)« + K4Fe(CN). = 2K,Zn,(Fe(CN)6),. 

* In testing for metallic poisons, chromic oxide (CrjOi), which is insoluble in 
adds, may be disregarded as it is not poisonous. 

• Two drops of ID per cent, potassium chromate solution ( = o .01 gram of 
KsCrOi) in 500 cc. of water produce a marked yellow color. Fifty cc. of this 
solution contain o .001 gram of KiCrOi which can still be recognized by the 
yellow color. 



I 



METALLIC POJSONS 163 

(o) Chrome Yellow Test.^Add acetic acid in excess to 
one portion of the filtrate, and boil for some time to expel 
carbon dioxide and nitrous acid as completely as possible. 
Then add a few drops of lead acetate solution. A yellow 
precipitate of lead chromate (PbCrO«, "Chrome Yellow") 
appears, if chromium is present. When the precipitate is 
mixed with considerable lead sulphate or chloride, the color 
is only yellowish. .A white precipitate is due to PbSO*, PbCU 
or PbaCPO^j. When the aqueous solution of the melt is color- 
less, such a precipitate is usually obtained. 

Potassium nitrate, always present in the melt, lead acetate and acetic Bcid> 
brought together in solution, produce a distinct yellow color, in which a 
white precipitate may appear yellow. To eliminate this source of error, 
allow the precipitate to setlle. collect upon a filter and wash thoroughly. H 
it is pure white, chromium is ahsect. 

(6) Rediiction Test. — Add sulphurous add to the second 
portion of the yellow filtrate. The yellow color changes to 
green, or greenish blue, with formation of chrome alum. This is 
not as delicate as the preceding test. 

METALLIC POISONS IV 

Detection of Barium, Lead and Silver iii the Residue from Hydrochloric 
Add and Potassium Chlorate 

Wash the residue left undissolved in the treatment with 

hydrochloric acid and potassium chlorate thoroughly with water 

and dry in the air closet or upon a porous plate. Then add 

three times the quantity of a mixture of 2 parts of potassium 

nitrate and i part of sodium carbonate and triturate in a mortar 

with the filter. Gradually introduce this nfixture into a hot 

porcelain crucible. In this operation organic substances (fats, 

fatty acids, etc.) are oxidized by potassium nitrate with con- 

|nderable deflagration. FinaUy when all the material is in the 

Icrucible, add 0.25-0,5 gram more of potassium nitrate. Cool 

' the melt, soften with water, wash into a flask and pass carbon 

dioxide through the turbid liquid for several minutes. This 

treatment converts caustic alkali into carbonate and completely 

[precipitates lead which may be in solution. Then heat the 



164 DETECTION OF POISONS 

solution to boiling and let settle for some time. Collect upon 
paper the sediment^ which may contain barium carbonate, basic 
lead carbonate and metallic silver. Silver gives the sediment 
a gray color. Thoroughly wash the precipitate with hot water 
and dissolve upon the paper in hot, rather dilute nitric add,' 
passing the acid through the paper several times. Evaporate 
this solution to dryness. Dissolve the residue in water, heat 
the entire solution to boiling, and precipitate silver with dilute 
hydrochloric acid. Filter, to remove silver chloride, and pass 
hydrogen sulphide through the filtrate to precipitate lead. 
To test for barium in the filtrate from lead sulphide, first boil 
to expel hydrogen sulphide and filter to remove insoluble 
matter. Then add dilute sulphuric acid which precipitates 
barium sulphate. 

To identify silver further, dry the hydrochloric acid precipi- 
tate and fuse in a porcelain crucible with a little potassium cya- 
nide. Extract the melt with hot water. Metallic silver re- 
mains undissolved. ^ 

To confirm the presence of lead, dissolve the hydrogen sul- 
phide precipitate in hot nitric acid and evaporate the solution 
to dryness. Dissolve the residue in water, filter and test the 
solution for lead with sulphuric acid or potassium chromate. 

To identify barium further, collect the sulphuric acid pre- 
cipitate upon paper, thoroughly wash and test upon a clean 
platinum wire in a non-luminous flame. Barium imparts a 
yellowish green color to the flame. To avoid any mistake, 
examine this flame with the spectroscope. These reactions 
of identification are always necessary in toxicological analysis. 

SYNOPSIS OF GROUP m 

Heat the residue from distillation of volatile poisons, or a 
portion of original material, in a glass flask or porcelain dish 

^ Even in the absence of barium, lead and silver, such a sediment nearly 
always appears. In that case it usually consists of the material of the porcelain 
crucible the glazing of which is partially attacked in the fusion process. 

* Use 5-6 cc. of an acid prepared by mixing i volume of concentrated nitric 
add and 2 volumes of water. 



METALLIC POISONS 



165 



upon the water-bath with dilute hydrochloric acid (12.5 per 
cent.) and potassium chlorate and shake frequently or stir. 
When most of the material is dissolved and the solution is 
yellow, dilute with water. Add a few drops of sulphuric acid 
and filter the cold solution. 



MateriaL Treated with HCl and KClOs. Dilute H1SO4. Filter. 



Filtrate.^ Saturated warm with HjS. 


Residue. Tested 
for "MetaUic 


Precipitate. Treated with hot 
(H4N),S, and (H4N)0H. 


Filtrate. Tested 
for "MetaUic 
Poisons III." 
Cr, Zn. 

• 


Poisons IV." 
Ag, Pb, Ba. 


Filtrate. Tested for 
"MetaUic Poi- 
sons I.'* 

As, Sb, Sn, Cu, 


Residue. Tested 
for "MetaUic 
Poisons II." 
Hg, Pb, Bi, Cu, 
Cd. 





Action of Heavy Metals 

Most salts of heavy metals, as lead, copper, mercury, silver, uranium and 
bismuth, precipitate proteins. These compounds are metaUic salts of albumins 
(albuminates). In combining with the oxides of these metals, proteins behave 
like acids. If the metal albuminates first formed are insoluble, or only sUghtly 
soluble in the body fluids, they are non- toxic or only slightly toxic. But a sol- 
uble albuminate is transported throughout the organism and exerts a toxic action. 
Every cell in contact with the dissolved metal may be poisoned. Mercury 
albuminate is an example of the latter class of metal albuminates. Being sol- 
uble in sodium chloride and protein solutions, it acts as a powerful poison. 
Copp>er albuminate on the other hand is not appreciably soluble in solutions of 
sodium chloride, hydrochloric acid or proteins. Not entering the circulation, it 
is as good as non-toxic. Lead and sUver albuminates are like copper albuminate 
as regards solubUity in the solvents mentioned. But if a heavy metal, which 
forms a difficultly soluble albuminate, finds its way into the organism in organic 
combination so that it cannot be precipitated by proteins, for example, copper in 
union with tartaric add, it then is as poisonous, or nearly as poisonous, as 
mercury in corrosive subUmate. Administration intravenously of 20 mg. 
of such copper causes the death of an adult rabbit. 



* If this filtrate contains much free hydrochloric acid, remove most of the 
acid by evaporation. Then add ammonia until alkaline and finally acidify 
with dUute nitric acid. 




166 



DETECTION OF POISONS 



Consequently precipitation takes place wherever the salt of a heavy metal 
comes in contact with proteins. The term corrosion is applied to such an 
occurrence. Tliere is always present the metallic oxide, protein and the add 
originally combined nith the metal. As a rule the add is loosely held by the pre- 
cipitate and is washed away by the circulating blood. The corrosive action of 
salts of heavy metals is due both to the union of the metallic oxide with protein, 
living protein being changed to dead metal albuminate, and to the caustic action 
ol the free add. Therefore the intensity of the action of the salt of the heavy 
metal depends upon the nature of the given metal albuminate. The degree of 
solubility is espedally important (see above), also the quantity and strength ot the 
free add. Salts of heavy metais not only may aSect the place of appUcation but 
they may give rise to serious changes where eliminated, as in the intestines or 
kidneys. Before elimination they may also seriously harm parenchymatous 
organs like the liver, as well as the drculatory organs. Finally salts of heavy 
metals have an important action upon the blood. R. Kobert and his collabor- 
ators have found that white as well as red blood-corpuscles may combine with 
metals and thus act as antidotes. Kobert has shown that the substance of red 
blood -corpuscles is capable of taking up a considerable quantity of a heavy metal. 
A chemical compound (metal himoglobb) is formed and the oxyhsmogtobin 
spectrum is not changed. Thus lead speedily impairs the vitality of red blood- 
corpuscles. Consequently red blood -corpusdes are lulled in large quantity in 
lead poisoning. 



Fate, Distribution and EliminatiOR of Metals in the Human Body 



I 



Arsenic.^Elimination of arsenic takes place mainly through the urine, 
begins several hours (7-13) after administration and, after a single dose, usually 
lasts 4-7 days. A great many experiments have shown the duration of arsenic 
elimination in urine to vary from a few days to several weeks. Some observers 
have found arsenic in urine So, and even go days, after poisoning. Conse- 
quently in suspected arsenic poisoning first 



« 



poisoning tht 
blood-corpusdi 

As regards 
are usuaUy found 



s usually diminished in volume and 



alburnin and 



enic by different organs, large quantities of the poison 
n the liver. Examine also the stomach and intestines with 
it ot the poison will obviously be in these organs in case of 
recent administration. The spleen, kidneys and muscles usuaUy contain arsenic. 
But the brain rardy shows more than traces of the poison. On the other hand, 
the bones in many instances have contained arsenic. This fact has given rise to 
the hypothesis that arsenic is capable of repladng phosphoric add in the bones. 
During the first stage in elimination of arsenic from the organism the poison 
appears to be deposited in the bones as caldurn arseniate. If large doses of 
difficultly soluble arsenic compounds, as while arsenic, or smaU doses of soluble 
compounds have been taken, the liver alone appears to arrest and retain the 
poison. Probably the quite stable arseno-nucleins are formed in that organ. 

From experiments where the organism was deluged with easily soluble arsenic 
compounds, Chittenden concluded that the brain can arrest and retair 



METALLIC POISONS 



167 



I ajid that arsenic then accumulates there. But after administration of irhite 
□ic OT Schweinfurt fcreen, only traces of the poison can be detected in the 
brain. Two instances of arsenical poisoning are in favor of this view. la the 
first case a woman died g hours after eating a highly arsenical soup. The liver 

I weighing 1159 grams, contained j5 mg, of arsenious oiide; the kidneys and 
bladder 0.6 mg.; and half the brain only a taint trace of the poison. In the 
Mcond case a young woman died a day after taking Schweinfurt green. In 

I tbis instance too the brain contained only traces of the poison. In experiments 

V upon animals most of the arsenic was always found in the liver. 



Normal Arsenic' 



The view that certain organs of the body may contain arsenic as an essential 
oonitituent has led to the use of the term "normal" to distinguish such arsenic 
from that entering the organism in food or drugs. The introduction of this term 
into chemical literature is unfortunaie, because it sujjgests the possibility of two 
kinds of arsenic Such a notion has no foundation whatever. Arsenic is arsenic 
and no test capable of showing more than one kind is known. A comnaittee of 
the French Academy of Sdences' after carefully investigating this matter came 
to the coadusion that arsenic never occurs normally ia the human body. But 
within recent years A. Gautier' after making many analyses of different materiab 
bas come to the opposite opinion. Gautier thus summarizes his results in one of 
llis papers:* 

"Speaking from a medico-legal point of view. I would state that arsenic, 
aade from the thyroid, mammary and thymus glands, never occurs in the human 
body except in the skin, hair, bones, mi|k and sometimes in the fxces and then 
only in traces which are often infinitesimal. Excepting the braia, the other organs 
and fluids, especially those forming the bulk of the body, as muscular tissue, liver, 
spleen, kidneys, lungs, blood, urine, etc., fail to show the slightest trace of arsenic 
. , If a chemist therefore examines individually these arsenic- 
free organs by my method or by one less delicate and finds traces, especially 
appreciable traces, of this metalloid, such arsenic has been absorbed during life 
dther medicinally or criminally." 

Gautier found the largest quantity of arsenic in the thyroid gland (0.75 mg, 
in 100 grams of gland) but this result has not yet been confirmed. Other 
chemists to be sure have found arsenic in the thj-roid gland but in much smaller 
qtiantity. These same chemists have also found arsenic in organs which Gautier 
says are noa-atsenicaL The following b a summary of their results but the 
original papers should be consulted for full details; 

I 'The brief account of "normal arsenic" in the German edition seems insuS- 
Pdent. After thoroughly examining the literature, the translator has therefore 
decided to treat this subject more fully. Tt. 

'Comptesrendusdel'Academiedes Sciences I >, 1076-iiog {t84i). 

•Ibid., tag, <)Z9~936 (1859). 

'Ibid. 130, »84-jgi (1900). 



^ 



168 



DETECTION OF POISONS 









'R«»nnii.rlr« AnH 


Observer 


Material 


Arsenic found 


conclusions 


Gautier' 


Human and ani- 


0.75 mgr. in 100 


Used 6 glands and 




mal organs and 


grams of human 


assumed uniform 




other material 


thyroid gland 


distribution of As 


Bertrand* 


Only animal ma- 


0.015 mgr. per 100 


Concludes As is a 




terial 


grams of dried 


normal constitu- 






sponge 


ent of protoplasm 


Schaefer' 


Human organs 


.007 mgr. per 100 


Concludes As may 






grams of human 


occur in all organs 






thyroid 


but found many 
free from As 


Pagel* 


Human and ani- 


Positive but not 


Found testes arseni- 




mal organs 


quantitative 


cal but Gautier 
says they are not 



On the contrary several chemists have carefully analyzed human and animal 
organs, either finding no arsenic or detecting this metalloid in mere traces, which 
are inconstant in occurrence and confined to no special organ. The table 
on the opposite page briefly summarizes their results. 

The results set forth in this table place "normal arsenic" in a doubtful 
position at least. If it is a reality and not a fancy, the quantity of arsenic, 
compared with that obtained in an analysis actually dealing with this metalloid, 
is so minute that the toxicologist need feel no concern. If he has conducted his 
analysis with every precaution as regards reagents and method and obtained a 
distinct mirror, he may dismiss the "normal arsenic" chimera and accept the 
result as due to arsenic that has entered the body from some external source. 
KunkeP has summed up the matter in these words. 

"The so-called normal arsenic, if there is such a thing, does not affect the 
results of forensic chemistry, because the so-called normal quantities are so 
exceedingly small (0.0 1 or even 0.00 1 mg. in an organ) that the quantities 
necessary to furnish a satisfactory forensic proof, which are a hundred or even 
a thousand times greater, must be regarded as an entirely different and much 
higher order of magnitudes." 

Antimony. — Elimination of antimony takes place largely through the urine. 
The rest of the metal is found chiefly in the liver and gastro-intestinal tract, as 
well as in the kidneys and brain. Pouchet found antimony in bones, skin, hair 

* Loc. cit. 

*Annalesde I'lnstitutPasteur 17,1-10(1903); Comptes rendus de TAcademie 
des Sciences 135, 809-812 (1902); and Bulletin de la Soci6t6 Chimique de Paris 
(3), 27, 1233-1236 (1902). 

* Annales de Chimie Analytique 12, 52-58 and 97-101 (1907). 

* Dissertation, University of Nancy, 1900. 

*Zeitschrift ftir physiologische Chemie 44, 511-529 (1905). 



METALLIC POISONS 



169 



Observer 


Material 


Results 


Conclusion 


HOdlmoser^ 


Human thyroid 


20 analyses 


Liver gives positive 




gland and liver 


Negative or few 


results as often as 






traces 


thyroid gland 


Cem^» 


Human and ani- 


28 analyses 


Minute traces may 




mal th3rroid and 


Negative or faint 


appear but not 




thymus glands. 


traces 


constant 




Human liver 






Ziemke* 


Various human 


Over 40 anal3rses. 


Not a normal con- 




organs 


Negative. One 


stituent of human 






case doubtful 


organs 


Wieser* 


Various human 


32 analyses. Mostly 


Traces inconstant 




and animal or- 


negative but few 


and due to chance 




gans 


traces 


contamination 


Kunkel* 


Various human 


Negative 


No such thing as 




and animal or- 




normal As 




gans 






Bloemendal* 


Various human 


Mostly negative 


No such thing as 




and animal or- 


but few traces 


normal As 




gans 






Warren' 


Human thyroid 


32 analyses. Nega- 


No such thing as 




gland 


tive except two 


normal As 






slight traces 





and chiefly in the intestinal tract. A large part of the ingested poison may be 
eliminated by emesis. A part is apparently retained in the body for some time, 
since antimony has been detected in the liver and in the bones months after the 
last administration. 

Lead. — ^Lead is eliminated in urine and faeces. Elimination by the feces 
always exceeds that by the urine, even when the lead has not been taken by 
the mouth. Mann^ (see R. Robert, Intoxikationen), for example, in the case 
of two patients was never able to find more than 0.6 milligram of lead in the 
urine collected during 24 hours, whereas the fseces during the same period con- 
tained 2-3 milligrams of lead. In lead poisoning the metal has been found in the 
saliva, bile and in both red and white blood-corpuscles. In animals relatively 

* Zeitschrift fUr physiologische Chemie 33, 329-344 (1901). 
*Ibid., 34, 408-416 (1902). 

' Vierteljahrsschrift fUr gerichtliche Medizin (3) 23, 51-60 (1902). 

* Dissertation, University of WUrzburg (1903). 
•Zeitschrift filr physiologische Chemie 44, 511-529 (1905). 

* Dissertation, University of Ley den (1908). 
' W. H. Warren, analyses not published. 

* Zeitschrift ftir physiologische Chemie 6, 6 (1882). 



170 DETECTION OF POISONS 

most of the lead has been found in the kidne3ns, after which come bones, liver, 
testes and finally the brain and blood. In experiments with she^ Ulenbeiger 
and Hofmeister obtained the following results: 

^ J ii . J Pb in grams per looo 

Organs and flmds 

grams 

Kidne3ns 0.44-0.47 

Liver 0.3 -0.6 

Pancreas 0.54 

Salivary glands 0.42 

Bile 0.11-0.40 

Bones 0.32 

Fseces 0.22 

Spleen o . 14 

Blood 0.05-0.12 

Urine 0.06-0.08 

Lead is eliminated especially by the bile and in acute poisoning this secretion 
may contain more lead than any of the other organs or secretions. Oliver^ has 
given the following results for human material: 

^ Pb in grams per 1000 

Organs 

grams 

Liver 0.0416 

Spleen 0.039 

Large brain 0.0216 

Small brain 0.0086 

Kidneys 0.013 

Heart o . 0005 

Elimination of lead by the urine is said, in the case of man, not always to be 
uniform. The urine is free from lead for a long time and later, without further 
administration, again contains the metal. This behavior is in harmony with the 
fact that lead can be retained in the organism many months. In chronic lead 
poisoning the brain has frequently been found to contain much lead. In this 
kind of poisoning elimination of the metal is always more abundant by the fsces 
than by the urine. 

Chromium. — Chromic add and soluble chromates and dichromates are quite 
toxic. The mucous membranes absorb alkaline chromates rapidly and severe, 
acute poisoning occurs. The poison causes intense pain in the stomach and 
intestines, collapse and kidney derangement which may terminate fatally in a 
few hours. Other symptoms are nausea, vomiting of yellow matter which later 
is tinged with blood, diarrhcea and even bloody stools, intense thirst, emaciation, 
great anxiety, severe pain in the abdomen, faint and quickened pulse — "the 
cholera picture.'' (Kimkel, Toxikologie.) The statements regarding the quan- 
tity of an alkaline chromate, capable of producing acute poisoning, agree fairly 
well. Even a few decigrams (0.2 gram) may cause very serious symptoms which 
sometimes terminate fatally. Chromic add is eliminated mainly by the urine but 

^ The Lancet, March, 1891. 



METALLIC POISONS 



Elinunation takes place rapidly and the body h 



I 



partly by the ir 

free of the poison. Four days after administration of quite large quantiliea ol a 

chromaie, the urine and tices are said to contain only traces of this metal. 

Copper. — Only a small amount of copper, varying with different compounds, is 
absorbed by the intestines and carried into the circulation. Sodium cupric 
tartrate and copper salts of fatly adds are absorbed most easily. Copper poison- 
ing rarely occurs from introducing a copper compound into the stomach. Copper 
compounds in large amounts act as local caustics and occasion severe pain La the 
stomach. Vomiting and the sense of taste make It impos^ble to take much of a 
copper compound. Foods containing copper are unpalatable. The sense ol 
taste as well as after- taste prevent one from swallowing such food in any quantity. 
Food, containing 0.5 gram of copper per kilogram, has a marked taste. Irresisti- 
ble nausea, steadily increasing, soon makes it impossible to take more of the food 
containing copjier. Elimination of copper by the urine is very slight. Copper 
absorbed from the intestines is arrested by the liver where it accumulates. 
Traces of copper have frequently been found in the human liver. The author in 
to»icological analyses has repeatedly found weighable quantities of copper in 
the livers of adults who had not taken copper salts beforehand, except possibly for 
suicidal purposes. The liver is the most important organ for the detection of 
copper, next to which come the bile, kidneys and the gastro-inlestinal mucosa. 
Copper is said to be in the liver as a nuclein compound. In the case of blood, 
copper is located not in the serum but in the corpuscles, 

Mercur;. — Distribution of mercury in the body is smd to be always the same, 
no matter what tbe method ot administration is. It is immaterial whether il is 
introduced by the mouth, hypodermically or from an abrasion. Elimination of 
mercury takes place through the saliva, sweat, bile, gastro-intestinal mucosa and 
urine. EUmination in the saliva seems to be constant, ^nce mercury can always 
be detected in the saliva during use of mercurials in lues. A relatively large 
quantity ot this metal is said to be eliminated in the sweat. Opinions differ as to 
the relative quantities eliminated by the urine and intestines. Usually elimina- 
tion by the intestines exceeds that by the kidneys. Recent experiments appear 
to show that mercury is ehminated in the urine regularly and in slowly increasing 
quantity and then slowly diminishes. Elimination of mercury ceases after 6-9 
months and even later. In a most favorable case the total quantity of mercury 
eliminated in the urine amounts to about 50 per cent, of the total quantity taken 
but is frequently much less. In mercury poisoning the kidneys, of all the organs 
pcTMStently contain most of the poison even for weeks. Then follow the liver, 
ipleen, bile and intestinal mucosa. In toxicological analysis the urine shoidd 
also be examined, though in acute poisoning it always contains only a fraction of a 

'"igram ot mercury in a Uter. In severe mercurial poisoning the metal may be 
to occur in all organs and sc 



Electrolytic Separatioii of Mercury from Urine 

Heat a liter ot urine upon the water-bath about 1 hours with 5-6 grams of 
potasauffi chlorate and to cc. of concentrated hydrochloric add and shake fre- 
quently. Evaporate to 300 cc. and use this solution for the electrolytic separa- 
tion of mercury. Use a Bunsen battery of 3-4 cells. Or any other galvanic appara- 



172 DETECTION OF POISONS 

tus having the same strength of current. A thin sheet or rod of gold 2 mm. thick 
and 6-10 cm. long serves as the cathode, and a piece of platinum wire of about the 
same thickness as the anode. Place the electrodes in the solution 2-4 cm. apart 
and allow the experiment to run 24-48 hours. Wash and dry the gold cathode, 
upon which the mercury is deposited, and place in a glass tube (20 cm. long and 4-5 
cm. in diameter). This tube is sealed at the bottom and reduced at the top to 
smaller size. Apply heat until all the mercury is expelled from the gold. De- 
posit the sublimate 3-4 cm. beyond the top of the gold rod. Then seal the tube 
below the sublimate. Introduce a small crystal of iodine into the tube and seal 
the other end. Heat the iodine carefully over a small flame to bring it into con- 
tact with the mercury. The two elements combine to form red mercuric iodide. 

Sometimes it is convenient to precipitate mercury from urine upon other 
metals, for example, copper, gold, brass and zinc dust. Witz heats to boiling 
with hydrochloric acid and concentrated potassium permanganate solution to 
destroy organic matter. Use 10 cc. of concentrated hydrochloric add and 15-30 
cc. of potassium permanganate solution for 500 cc. of urine. Slowly pass the 
decolorized liquid through a glass tube over a copper spiral. Wash the copper 
with potassium hydroxide solution, then with absolute alcohol and cleanse with 
filter paper. Finally dry at 70-80** and heat in a glass tube. Treat the mercury 
sublimate with iodine as described. Satisfactory deposition of mercury upon 
other metals (gold, copper) requires previous destruction of organic matter in the 
urine by hydrochloric acid and potassium chlorate. Otherwise, organic substances 
deposited upon the metal interfere with the iodine test for mercury. 

Silver. — In severe poisoning silver has been found in bile, fseces and in many 
organs. In acute poisoning the urine is usually free from silver, whereas the 
contents of the intestines may contain the metal even after subcutaneous injec- 
tion. Absorbed silver salts appear to be reduced in all parts of the body. Ex- 
amination of the bodies of persons who have suffered from argyria (chronic silver 
poisoning) has disclosed precipitation of metallic silver in the organs. Silver 
salts, added to albumin solutions, form very stable compounds usually amor- 
phous. Silver may evince even greater affinity for albumin than for chlorine. 
All observers agree that only very little of the silver reaching the intestines is 
absorbed. After silver medication the stools are black from silver sulphide. 
Frequent quantitative estimations of silver in argyrotic organs have been made. 
In one case the liver gave 0.047 per cent, of silver and the kidneys 0.061 per cent. 
In the condition called argyria, or argyrosis, the skin is black. Internal adminis- 
tration of silver salts causes this color to develop gradually. By degrees it may 
become quite marked even causing disfigurement. 

Qtxantitative Estimation of Silver in Organs and Urine 

V. Lehmann,* in determining silver in organs (liver, kidneys, spleen, brain), 
first thoroughly dries the finely ground material and then fuses with sodium 
carbonate and potassium nitrate. Extract the melt with water and dissolve 
the insoluble residue of metallic silver in hot nitric acid. Evaporate this solution 
to dryness up>on the water-bath and precipitate silver as chloride. Avoid a 
large excess of hydrochloric acid. 

* Zeitschrift fUr physiologische Chemie 6, 19 (1882). 



I METALLIC POISONS 173 

Mix urine with sodium carbonate and potassium nitrate, evaporate to dryness, 
fuse the residue and treat the melt as described. 

nranitmi. — Experiments made by R. Kobert have shown that uranium, 
administered subcutaneously or intravenously, is the most Ionic of all melals. 
Uranyl acetate is an excellent precipitant of albumins and the other uranyl salts 
must behave in much ihe same way. Consequently internal administration 
of concentrated solutions of uranyl sails destroys the mucous surfaces they 
touch, tor example, that of the stomach, changing the living stomach wall lo 
dead uranyl- albuminate. Uranyl salts therefore must be classed among the 
poweri'ul caustic poisons. In addition to acting as local corrosives, uranium 
salts resemble hydrocyanic acid in partially arresting internal oxidation in the 
organs and occasioning the severest disturbances of metabolism. 

Biamuth. — This metal becomes quite toxic when it reaches the blood. Bis- 
muth solutions, prepared by dissolving bismuthous hydroxide in tartaric or citric 
acid and then neutralizing with sodium or ammonium hydroxide solution, have 
been repeatedly administered lo animals subcutaneously and intravenously. 
The smallest lethal dose of these double bismuth salts, injected subcutaneously, 
was found to be but 6 mg. per kilogram for a dog or cat and 14 mg. per kilo- 
gram (or a rabbit. Bismuth salts insoluble in water produce entirely differ- 
ent results when admimstered internally. Bismuth subnitrate and similar 
salts dissolve very slightly in the highly diluted hydrochloric add of the gastric 
juice. Consequently very little bismuth is conveyed to the blood. Most of the 
bismuth taken by the mouth reaches the intestines. Instead of being absorbed, 
it is chatiged to bismuthous sulphide by the hydrogen sulphide always present. 
Absorbed bismuth is eliminated by the saliva, bile, urine, mucous lining of the 
moulh, stomach, small and large intestine and also the milk. If an animal is 
ptHsoned by bismuth, the metal can be detected in the uriae, bile, liver, kidneys, 
spleen, walls of the intestines as well as in the bones. Different observers have 
found the metal in especially large amount in the milk but very little in the kid- 
neys and liver. 

Snc. — There is no doubt thai zinc salts reachirvg Ihe intestinal tract are 
absorbed in very small quantity. .\s yet there is no satisfactory explanation of 
the fale of absorbed zinc. In zinc poisoning large amounts of the metal have been 
found repeatedly in the liver and bile. This may mean that zinc is arrested by 
the liver and eliminated in the bile. Lehmann' after 335 days killed a dog that 
had been fed for a considerable lime upon zinc carbonate. The following organs, 
arranged according to the quantity of metal in each, contained xinc: liver, bile, 
large inlesline, thyroid gland, spleen, pancreas, urine, kidneys, bladder, muscle, 
brain, lymphatic gland, stomach, small intestine, lungs, blood. OccaMonally 
considerable quantities of zinc may be taken with articles of food and drink. 
All adds dissolve metallic zinc very freely. Even water containing carbon diox- 
ide is a solvent. Consequently it may be in drinking water from galvanized 
pipes. All kinds of food and drink, kept in zinc vessels or vessels coated with 
zinc, may contain more or less of this metal. Plants grown upon soil containing 
LsIdc take up the metal. Zinc has also been found repeatedly in parts of human 
^■tdavers under circumstances precluding all possibility of poisoning by this 

■ ' Archiv fur Hygiene j8, 291 (iSgS), 



174 DETECTION OF POISONS 

metal. Even considerable quantities of zinc have been found in the human 
liver. 

Tin. — The cases of tin poisoning thus far observed resemble those of copper 
and zinc. What knowledge there is regarding the toxic action of absorbed tin 
has been gained from experiments upon animals. These experiments show that 
small quantities of tin are absorbed and eliminated in the urine, when ordinary 
tin compounds are brought into the stomach. But thus far distinct symptoms 
of poisoning by such quantities of the metal have not been confirmed. (Kunkd, 
Toxikologie.) 

White^ failed to produce poisoning by bringing tin into a dog's stomach. The 
animal received sodium stannous tartrate in increasing doses for 22 days, the 
daily amount being 0.02-0.06 gram. Yet the animal absorbed tin. In the urine, 
during an experiment lasting 8 days, White found 0.02 gram of tin. But the 
tin salt mentioned, introduced directly into the circulation of the animal, was 
quite toxic in its action. Stannous chloride, administered for a very long time 
to a dog, produced symptoms of poisoning. The urine in this case contained 
small quantities of tin. Kunkel (Toxikologie) states that tin has a very slight 
poisonous action. Apparently it is eliminated very rapidly by the kidneys. 
Quite probably this prevents accumulation of the metal in the body and conse- 
quent poisoning. The fact that White did not observe toxic symptoms, after 
feeding a dog for 22 days with relatively large quantities of easily absorbable 
sodium stannous tartrate; and that Ungar and Bodlander' failed to produce 
derangements with the same compound, until it had been administered for a year, 
prove that tin is quite free from toxic properties. Hence, tin vessels may be used 
and preserved articles of food containing tin have practically no deleterious 
action upon health. 

^ Archiv f(ir experimen telle Pathologie und Pharmakologie 13, 53. 
• Zeitschrift f(lr Hygiene 2, 241. 



CHAPTER IV 

POISONS NOT IN THE THREE MAIN GROUPS 

MINERAL ACmS 

Hydrochloric, Nitric and Sulphuric Adds 

To detect free mineral acid, extract a portion of material with 
cold water, filter and test as follows, if the solution is strongly 
add: 

1. Methyl Violet Test — ^Add a few drops of an aqueous 
(o.i :iooo) or alcoholic (i :ioo) solution of methyl violet^ to 
a small portion of filtrate. A free mineral acid produces a blue 
or green color. 

2. Methyl Orange Test — Add a few drops of a dilute aqueous 
solution of methyl orange^ to the filtrate. A red color indicates 
free mineral acid. 

3. Congo Paper Test. — Even very dilute solutions of free 
mineral acids turn "Congo paper'' blue. 

4. Giinzburg's Test — Mix a few drops of the filtrate with 
3-4 drops of Giinzburg's reagent' and evaporate to complete 
dryness upon the water-bath, or over a small flame. Free 
hydrochloric or sulphuric acid gives a fine red or reddish yellow 
residue. Nitric acid gives more of a yellowish red residue. 

* Methyl violet is the hydrochloride of hexa-methyl-para-rosaniline: 

(CH,),N.C6H4v /CH = CHv V 

>C = C< >C = N(CH,),C1 (quinoidal form) 



or 



(CH,),N.C,H4^ \CH= CH' 



V 



(CH,),N.CeH4v /C,H4. N(CH,),C1 
(CH,),N.C.H4' 






• Methyl orange = Dimethyl-ainino-azobenzene-4-sulphonic acid: 

4 II 4 

(CHi), N.CtH4.N-N.CeH4.SO,OH. 

The sodium salt of this sulphonic acid also appears in commerce under the name 

"methyl orange." 

* See page 314 for the preparation of this reagent. 

176 



176 DETECTION OF POISONS 

5. Sn^iliocyaiiate Test — Add a little potassium sulphocya- 
nate solution to ferric acetate solution and dilute with water 
until vellow. Then add the solution to be tested. Free mineral 
acid produces a* blood red color. Traces of free mineral add, 
especially if considerably diluted, do not give a red color until 
se\*eral minutes have elapsed. 

One or more of these general tests, which furnish evidence of a 
free mineral acid, must always accompany the special tests to 
be described later. Not only free mineral acids give the special 
tests but in certain cases their salts. Chlorides, sulphates and 
nitrates are normal constituents of nearly all vegetable and 
animal materials. As a rule an examination of cadaveric 
material for mineral acids is necessary only when the autopsy 
jxMnts conclusively to such poisoning. That is to say, when 
ihow* are characteristic corrosions and discolorations about the 
Uoo. mouth, oesophagus and stomach. If general tests show the 
presence of free mineral acid, make special tests for the particu- 
lar acid. 

Hydrochloric Add 

l^ Chlorine Test. — Warm a little of the aqueous extract, if 
>hs'4 t^v dilute, with finely powdered manganese dioxide. Free 
V\;t\vhK>ric acid yields chlorine, recognized by its color and 
\NK>r. or by passing the gas into potassium iodide solution and 
'^tv^Atu^ iodine. Hydrochloric acid exclusively does not give 
t.\»^v u^sl. A chloride (NaCl) and free sulphuric acid give chlor- 
'Uv v^Wvler the same conditions. 

|y DtetillAtion. — If possible, separate hydrochloric acid from 
vs^v* Mibstances by distillation. The concentration of the 
u 4\l tx e^^Hvially important, since dilute hydrochloric acid upon 
«UauIU\U\M\ ttt first yields only water. Hydrochloric acid^ 

• U\U vlvv«i not begin to distil until the concentration is about 

• v^ 'V* vvnt. Since a dilute hydrochloric acid is usually ex- 
k\uaK\li vUstil the material mixed with water, or preferably a 

' li% «iw vU^lilUtlon of 100 cc. of i per cent, hydrochloric acid, the first 90 cc. 
»:**utUiv ^iU vx>ntttin only traces of hydrochloric acid, whereas the last portion 
x« .1; xxM»uiw WH^t of the acid. 



POISONS NOT IN THE THREE MAIN GROUPS 177 

filtered aqueous extract, nearly to dryness. In such a dis- 
tillation apply heat by means of an oil bath. To detect hydro- 
chloric add in the distillate, acidify with dilute nitric acid and 
add silver nitrate solution. Frequently a quantitative estima- 
tion of hydrochloric acid is required. In the absence of other 
acids, titrate the distillate with o.i n-potassium hydroxide 
solution, using phenolphthalein as indicator. Otherwise, esti- 
mate the acid gravimetrically, precipitating with silver nitrate 
and weighing silver chloride, or volumetrically by Volhard's 
method. In the latter case, precipitate hydrochloric acid with 
O.I n-silver nitrate solution in excess and subsequently estimate 
that excess by titration with o.i n-ammonium sulphocyanate 
solution, using ferric alum as indicator. Since the human 
stomach normally contains 0.1-0.6 per cent, of free hydro- 
chloric acid, an examination of stomach contents for this acid 
must always include a quantitative estimation. 

Nitric Add 

The human body normally contains only a very small amount of nitrates. 
When present, they are due usually to vegetable foods which contain small 
quantities of nitrates. Human urine almost always shows traces of the salts of 
nitric and nitrous acids. The chemical examination of cadaveric material need 
not include tests for nitric acid, unless the autopsy affords evidence of poisoning 
by this acid, as distinct signs of corrosion about the lips, mouth, oesophagus and 
stomach and sometimes perforation. These parts are more or less yellow or 
yellowish brown. A yellow froth is said to exude from the mouth and nose of 
the cadaver. Also the stomach contents are yellow in concentrated nitric add 
poisoning. If the concentration of the acid is less than 20 per cent., these specific 
changes may not appear in the gastro-intestinal tract. Nitric acid taken inter- 
nally, dilute or concentrated, appears at once in the urine. 

Detection of Nitric Add 

I. Distillation. — If possible, extract the material direct with 
water, filter and test the filtrate for nitric acid in the usual way. 
When the quantity of nitric acid is large, separate it from other 
substances by distilling the filtered aqueous extract. Apply 
heat by means of an oil bath. Nitric acid^ does not distil, until 

^ If 100 cc. of I per cent, nitric acid are mixed with bread crumbs and distilled, 
most of the acid will be in the final 10 cc. of distillate. 
12 



178 DETECTION OF POISONS 

it reaches a definite concentration. At the same time a large 
part of the acid combines with organic substances, if any are 
present, forming nitro-derivatives, xanthoproteic acid, etc. 
Nitric acid may also cause oxidation. Consequently the dis- 
tillate does not contain all the acid originally present. The 
residue from. such a distillation is usually distinctly yellow. 
Toward the end]of distillation brown vapors of nitrogen peroxide 
often appear. Such a distillate, added to starch paste and po- 
tassium^iodide, produces an immediate blue color in presence of 
dilute sulphuric acid. 

To detect nitric acid in the distillate, employ the following 
tests: 

2. G. Reuiy's Procedure.' — Extract the finely divided mate- 
rial with absolute alcohol, filter and add slaked lime in excess 
to the filtrate. To decompose any nitric acid ester present, let 
the mixture stand 12 hours, filter and evaporate the filtrate to 
dryness. Dissolve the residue in 95 per cent, alcohol, expel 
alcohol from the filtered solution and finally test an aqueous 
solution of the residue for nitric acid. Fleury has obtained by 
means of this method about 20 per cent, of the nitric acid from 
animal material. This procedure converts the acid into its 
calcium salt which is soluble in alcohol. But sodium nitrate 
is also quite soluble in 95 per cent, alcohol (i 150). There- 
fore, if the final residue gives a faint test for nitric acid, the proof 
of free acid in the original material is not conclusive. The 
following method obviates this diificulty. 

3. Baumert's Procedure.^ — Neutralize the material itself, or 
its aqueous extract, with milk of lime, dry and extract with 
alcohol. Or, after neutralization with milk of lime or calciimi 
carbonate, evaporate to a syrup and mix the latter while stir- 
ring with alcohol. Distil the filtered alcoholic extract obtained 
in either way, dissolve the residue in water, filter and evapo- 
rate the solution. Dissolve the residue again in alcohol and 
allow this solution to stand for several hours in a closed flask 
with about the same volume of ether. Filter this alcohol- 

^ Annales de Chimie analytique appliqu6e 6, 12. 
'Lehrbuch der gerichtlichen Chemie, second edition (1907). 



POISONS NOT IN THE THJfEE MAIN GROUPS 



179 



I 



ether solution, evaporate the solvent and dissolve the residue 
in a little water. Apply the following nitric acid tests to this 
solution : 

, (a) Diphenylamine and Sulphuric Acid Test.^B!ue color. 
[ Add a few drops of diphenylamine sulphate solution^ to the 
aqueous extract, or distillate, and carefully pour this nii.xture 
upon pure concentrated sulphuric acid free from nitric acid. 
If nitric acid is present, a blue zone appears where the two liquids 
meet, 

(h) Brucine and Sulphuric Acid Test. — Red color. 

MLx the liquid to be tested with the same volume of brucine 
sulphate solution^ and carefully pour this mixture upon pure 
concentrated sulphuric acid. If nitric acid is present, a red 
zone appears where the two liquids meet. 

(c) Ferrous Sulphate and SiUphuricAcidTest^Saturate the 
liquid to be tested with pure ferrous sulphate and carefully 
pour tliis solution upon pure concentrated sulphuric acid. If 
nitric acid is present, a black zone appears where the two liquids 
meet. 

(d) Copper Test. — Place a small piece of clean copper fwire 
or sheet) in nitric acid and heat. Red-brown vapors of nitrogen 
peroxide (NO 2) appear. 



Sulphuric Acid 



Nearly all animal and vegetable substances normally contain sulphates. Con- 
sequently an examination for free sulphuric acid must e:ic!udc its salts. There 
is no need of examining cadaveric material for the free add, unless marked corro- 
sion and discoloration of lips, mouth, cesophagus and stomach indicate its presence. 
There are eschars upon the Ups and the mucous lining of the mouth is grayish 
white. The white coatittg on the back of the tongue may have been dissolved 
exposing the lirm, brownish muscular tissue beneath. The tongue often looks 

J* Prepare this solution by dissolving 1 gram of diphenylamine, (C*H,)!NH 
B5 grams of dilute sulphuric add and too cc, of water. 

' Prepare this solution by dissolving 1 gram of bnicine in 5 grams of dilute 
sulphuric add and 100 cc. of water. The sulphuric acid used must give none of 
the tests for nitric acid. If it does not meet this requirement, heat in a platinum 

dish to expel interferitig nitrous substances. Or distil the add from a small 

^Hwtort. rejecting the first part of the distillate. 



i 



180 DETECTION OF POISONS 

as if it had been boiled. The mucous lining of the oesophagus is much wrinkled 
and coated gray. Externally the stomach is usually brown or slate-gray and its 
contents black. Frequently in sulphuric add posioning there is perforation of 
the stomach wall and brownish black masses find their way into the abdominal 
cavity. There may be black spots in the stomach, due according to R. Robert 
(Intoxikationen) not to charring, as previously supposed, but to brown-black 
haematin. Acids decompose the blood-pigment oxyhemoglobin mainly into 
haematin and protein (globidin). Methsemoglobin and haematoporphyrin may 
also be formed. Acids produce the latter from haematin and in the change there 
is loss of iron. All three of these decomposition products of the red blood-pig- 
ment, namely, methaemoglobin, hsematin and haematoporph3nin may be formed 
successively and then appear in the urine. The blood in the stomach walls is 
often acid and then contains chiefly methaemoglobin and hsematin. The mucosa 
of the intestines even far down may be grayish white and strongly acid. 



Detection of Sulphuric Acid 

1. Extract the finely divided material, if strongly acid, with 
cold absolute alcohol and after some time filter. The solution 
contains sulphuric acid but not sulphates. Evaporate the 
alcoholic filtrate upon the water-bath, or, if the volume is large, 
distil the alcohol. Dissolve the residue in a little water (lo cc.) 
and heat the solution to boiling to saponify^ ethyl sulphuric 
acid. Filter and test the filtrate with barium chloride or lead 
acetate solution. To prove that the precipitate is a sulphate, 
mix with sodium carbonate and fuse upon charcoal. The 
sodium sulphide formed blackens metallic silver in presence of 
water, or gives hydrogen sulphide with acids. 

2. Extract the finely divided material with water and apply 
the following tests to the filtrate: 

(a) Sugar Test. — Evaporate some of the filtered extract in a 
porcelain dish with a small particle of sugar. Free sulphuric 
acid produces a black, carbonaceous residue. 

(b) Sxilphur Dioxide Test. — Concentrate the filtered extract 
upon the water bath and heat in a test-tube with a few pieces of 
copper. Free sulphuric acid generates sulphur dioxide, recog- 
nized by its stifling odor. Distil the sulphur dioxide (preferably 
in an atmosphere of carbon dioxide) into a little water and test 
the distillate as follows: 

1 HO.SOi.OCiHs -h H,0 = C,Hs.OH -f HiSO*. 



POISONS NOT IN THE THREE MAIN GROUPS 181 

a. Warm some of the liquid with a little stamious chloride 
solution. A yellow precipitate of stannic sulphide* appears. 

0. Add iodo-potassium iodide solution drop by drop. The 
color of the iodine disappears and at the same time sulphuric 
add is formed: 

H,SO. + H,0 -h It = H,S04 -h 2HI. 

Barium chloride then precipitates barium sulphate insoluble 
in dilute hydrochloric acid. 

To estimate sulphuric acid quantitatively, either precipitate 
and weigh barium sulphate in the usual way, or titrate with 
o . I n-potassium hydroxide solution, using phenolphthalein as 
indicator. 

1000 cc. of o. I n-potassium hydroxide solution = 0.1 gram- 
equivalent of sulphuric acid = 4.9 grams of H2SO4. 

Detection of Sulphurous Acid 

Sulphur dioxide acts most injuriously when inhaled. It is very irritating to 
the respiratory organs and also changes the blood-pigment. After death the 
respiratory organs are found to be profoundly altered as when acted upon by 
strong mineral adds. After severe poisoning by vapors containing sulphur 
dioxide, the blood is dirty brownish red' and usually gives the hsematin spectrum. 
Human beings experience discomfort, if there are 0.015-0.02 volumes of sulphur 
dioxide per 1000 volumes of air. Many persons become quite ill in a few minutes, 
when there are 0.03 volumes of sulphur dioxide in 1000 volumes of air. The gas 
produces a sharp, stinging sensation in the nostrils, sneezing and coughing. In 
experiments upon mice, rabbits and guinea-pigs, Lehmann observed marked 
toxic symptoms from air containing 0.04 volume per cent, of sulphur dioxide; 
death ensued in 6 hours from 0.06 per cent.; and in 20 minutes from 0.08 per cent. 
Articles of food and drink, preserved by means of sulphurous acid or its salts, 
may injure the health, causing especially gastro-intestinal catarrh and other 
chronic derangements. For this reason it is prohibited to preserve articles of 
food and drink by means of sulphurous acid, sulphites and hyposulphites. 

If the quantity of sulphur dioxide in air is not too small, its presence may be 

^ Sulphurous acid and sodium sulphite, added to stannous chloride solution 
not too strongly acid, precipitate stannous sulphite, SnSOi, white and readily 
soluble in hydrochloric acid. Warmed in presence of hydrochloric acid, sulphur 
dioxide acts upon a stannous salt as an oxidizing agent. A precipitate of SniOioSs 
is formed, or H2S is evolved and SnCU formed, depending upon the amount of 
hydrochloric acid present. (Prescott and Johnson, Qualitative Chemical Analy- 
sis. Fifth edition, page 86.) 

' Neutral sulphites cause the blood to become brick red. 



182 DETECTION OF POISONS 

recognized by its characteristic stifling odor. A strip of paper, moistened with 
a solution of pure potassium iodate (KIOi) and starch, turns blue in air contain- 
ing sulphur dioxide owing to the formation of a compound of iodine and starch. 
This reaction serves as a preliminary test for the detection of sulphurous add and 
hyposulphites in chopped meat, sausage meat and other meat products. Shake 
the meat in an Erlenmeyer flask with phosphoric add, suspend in the neck of the 
flask from the stopper (see Fig. i, page 3) a paper strip prepared as described 
and heat the flask upon the water-bath. The paper should not turn blue. 

Explanation. — Sulphur dioxide reduces potassium iodate (a). Sulphuric add 
thus formed liberates hydriodic and iodic adds from their salts (fi andy). The 
iodine set free by the interaction of these two adds (5) finally turns the starch 

blue. 

(a) KID, -h 3H1SO, - KI +3H,S04. 

(/3) 2KI + H,S04 - 2HI + K,S04, 

(y) 2KIO, 4- HiSOi = 2HIO, 4- K,S04, 

(«) mo, + sHi =31, +3H,0. 

The offidal directions^ for the detection and quantitative estimation of sulphur 
dioxide in meat are as follows. Mix 30 grams of findy chopped meat with 200 
CO. of boiled water in a 500 cc. distilling flask.' Add sodium carbonate solution 
until the reaction is faintly alkaline. 

Let the mixture stand for an hour and then completely expd air from the ap- 
paratus by passing carbon dioxide through the tube extending to the bottom of 
the flask. Then introduce into the Peligot tube (see below) 50 cc. of iodine 
solution (5 grams of pure iodine and 7 . 5 grams of potassium iodide in a liter 
of water). Raise the stopper of the distilling flask and, without stopping the flow 
of carbon dioxide, add 10 cc. of 10 per cent, phosphoric add solution. Then 
carefully heat the contents of the flask and distil half the liquid, maintaining all 
the while a current of carbon dioxide. Transfer the contents of the Peligot 
tube, which should be brown, to a beaker, rinsing it out with water to prevent 
loss of solution. Add a little hydrochloric acid, heat and by means of barium 
chloride solution completely predpitate the sulphuric acid formed from the oxida- 
tion of sulphurous acid by iodine. 

HiSOi 4- H,0 4- It = H,S04 4- 2HI. 

If this test is positive, then the meat examined contains dther free sulphurous 
acid, sulphites or hyposulphites. In the quantitative estimation the barium 
sulphate should be weighed in the usual manner. 

OXALIC Acm 

Oxalic add and its salts, for example, salt of sorrel, are quite toxic substances. 
Administration of oxalic add has terminated fatally in the case of adults in a few 

1 Measures for putting into effect the law of the German Empire of June 3, 
1900, relating to the inspection of beef-cattle and meats. 

* The apparatus prescribed for ofiidal examinations is a distilling flask, having 
a capadty of 400-500 cc. and provided with a two-hole stopper for two glass 
tubes entering the flask. One tube extends to the bottom of the flask and the 
other only into the neck. The latter is connected with a Liebig condenser to 
which a Peligot tube is fastened at the other end by a tight stopper. 




POISONS NOT IN THE THREE MAIN GHOTTPS 

r.inlnutes. Oxalic add is veiy abundant in the vegetable kingdom in tbe form of 
i acid potassium salt. KHCjOi, and calcium salt. Eoircl, wood-sorrel and 
rhubub are especially rich in salts of oxalic add. Hence this add may find 
access to the body through food and drugs of vegetable origin. Moreover, 
oxalic add is a normal constituent in small quantity of human urine, 2-6 milH- 
grams being excreted in the course of a day. Consequently ic 
material it is often necessary to supplement a positive qualitative test by a 
quantitative estimation of oxalic add. 

To^ Action.- — An important difference between mineral adds and oxalic 
add is the toiidty of salts of the latter. Not only do free oxalic add and its 
add potas^um salt, salt of sorrel, show poisonous properties but even very dilute 
solutions of neutral sodium oxalate, NaiCiO,, act in the same way. Therefore 
in oxalic add poisoning it is necessary to distinguish between local corrosion, 
occurring at the point of application and also in part upon elimination, and re- 
mote action due to absorption. Local action at the point of application is cot- 
roave like that of all adds. Local action at tbe place of elimination depends upon 
the formation and insolubility of caldum oxalate. On account of the ease with 
which the organism takes up oxalic acid and its alkah salts, the action of the ab- 
sorbed poison is rapid. The effects caused by its presence may be attributed 
to tbe fact that this add removes in part from organs, as the heart, and from 
body fluids (blood) the caldum they require tor their life processes, converting 
it in part into insoluble caldum oxalate. Oxalates diminish the coagulating 
power as well as the alkalinity of blood. On the other hand they increase the 
[.quantity of sugar in the blood. In oxalic add poisoning there is a depression 
K,<if the entire metabolism. This b also the case as regards taking up oxygen and 
Fjllving off carbon dioxide. The body temperature faUs as the processes of metabo- 
lism are retarded. Owing to withdrawal of caldum from the heart, that organ 
is weakened and finally paralyzed. I.^cal action upon the kidneys is due to 
dogging of the injured urinary tubules by depoata of caldum oxalate. The flow 
of urine may wholly cease in consequmce of total impairment of the urinary 
tubules and death may ensue from anuria and urzmia. Fatal poisonings from 
large doses of oxalic add are usually of short duration. R. Kobert (Intoxtka- 
tionen) describes a case where death occurred within 10 minutes. 

Bischoff' has made statements in regard to the distribution of oxalic add in 
the different organs of persons poisoned by this substance. In a case, which ter- 
minated fatally in less than 15 minutes, the quantity of oxaUc add in each organ 
was determined separately and found to be: 



Weight 


Organ 


Oxalic Add 


1140 grams 


Stomach, oesophagus, intestine and 






contents 


1 . 2& grams. 


7;o grams 


Liver 


. a8s grams. 


I go grams 


Kidneys 


0.0:4s grams. 


180 grams 


Blood from the heart 


O.D43S grams. 


40 grams 


Urine 


0.0076 grams. 



'Berichle der Deulschen chemischen Gesellschaft, t6, 1350 (i88j). 



184 



DETECTION OF POISONS 



The quantity of oxalic add in the liver is noticeably large. The kidneys and 
urine contain only a little of the poison, owing to the short duration of life after 
poisomng. A striking thing about the urine excreted during oxalic add poisoning 
is the abundant deposition of crystallized caldum oxalate. 



Detection of OzaHc Add 

To detect oxalate without discriminating between the free 
acid, acid potassium salt or calcium oxalate, employ the follow- 
ing method: 

Add to the finely divided material 3-4 volumes of alcohol 
and acidify with dilute hydrochloric acid. Stir frequently and 
let the mixture digest 1-2 hours cold. Then filter through a 
plaited paper moistened with alcohol and wash the residue with 
alcohol. To prevent formation of ethyl oxalate during evapo- 
ration, add about 20 cc. of water to the total filtrate. Evapo- 
rate upon the water-bath until all alcohol is expelled. Pass 
the aqueous residue through a small filter. Extract the filtrate 
in a separating funnel 3-4 times with 50-60 cc. portions of ether. 

Let the total ether extract stand for 
some time in a dry fiask, then pass 
through a dry filter and distil. Dis- 
solve the residue in 2-3 cc. of water 
and pass the solution, if necessary, 
through a moist filter. Add am- 
monium hydroxide solution until 
alkaline and then saturated calcium 
sulphate solution. If there is a pre- 
cipitate, acidify with acetic acid and 
let solution and precipitate stand 
over night in a covered beaker. If 
there is still a crystalline precipitate, it can be only calcium 
oxalate. A microscopic examination of this precipitate is 
advisable. Calcium oxalate forms characteristic octahedrons 
having the so-called envelope shape (Fig. 17). When thor- 
oughly washed, calcium oxalate may be converted by ignition 
into calcium oxide which may be weighed. 

CaO : HjCa04.2HiO = Wdght of CaO : x 
(56) (126) found 




Fig. 17. — Caldum Oxalate 
Crystals. 



POISONS NOT IN THE THREE MAIN GROUPS 185 

Calcnlatioa. — Since the quotient 56 : 126 *■ 0.444, multiply the weight of 
caldum oxide found by 0.444 to get the corresponding amount of aystallized 
oxalic add. 



FREE ALKilLIES 

Potawriiim, Sodium and Ammoniuiii Hydroxides 

Free ADcalies. — ^The same general prindples used in detecting mineral adds 
are applicable also to the alkalies. Since potassiimi and sodium compoimds are 
normal constituents of animal and vegetable organisms, and since ammonia is a 
decomposition product of nitrogenous organic matter, the examination must 
alwa3rs show that the alkalies are in the free state, for they alone and their car- 
bonic add salts decompose and corrode animal tissues and not their neutral salts. 

Poisonings due to caustic alkalies resemble those caused by corrosive adds. 
If taken internally, their corrosive action gives rise to pain in the mouth, throat, 
oesophagus, stomach and abdomen. Mineral add corrosions are dry and brittle, 
whereas those from caustic alkalies are soft and greasy. The alkaU albuminates 
formed become gelatinous, swell and may partly dissolve in presence of much 
water. The destructive action of the caustic alkalies extends deep and affects 
the parts around the corroded places. In caustic alkaline solutions gelatinous 
tissues, homy substances, hair and skin swell considerably and finally dissolve. 
The stomach in alkali poisoning is softened in places, corroded and deddedly 
bright red in color. 

Detection of Alkalies 
Ammonia 

Free ammonia is usually recognized by its odor. A piece of 
moist red litmus paper, held over the material, becomes blue. 
A paper moistened with mercurous nitrate solution is blackened. 

Distillation. — If the material is strongly alkaline, extract 
several times with absolute alcohol. Use a flask with a glass 
stopper and distil the combined extracts. Collect the distillate 
in a little dilute hydrochloric acid and evaporate the solution 
in a porcelain dish to dryness upon the water-bath. Dissolve 
the residue in water and test the solution for ammonia, using 
Nessler's reagent and chloroplatinic acid. 

Fixed Alkalies 

The residue from the above distillation may contain potas- 
sium and sodium hydroxides. If the residue is strongly alka- 
line, first add a few drops of phenolphthalein and then excess of 



186 DETECTION OF POISONS 

barium chloride solution. The red color and the alkaline reac- 
tion, if due to carbonates, disappear, because two neutral salts 
are formed: • 

K,CO, -f BaCl, = BaCO, -f 2KCI. 

But if alkaline hydroxides are present, the alkaline reaction 
and red color remain, for soluble barium hydroxide is formed: 

2KOH -f BaCl, = Ba(OH), -f 2Ka. 

And the solution of this compound reddens phenolphthalein. 

To distinguish potassimn from sodium hydroxide, neutralize 
the residue from distillation with dilute hydrochloric acid and 
test for potassium and sodium as follows: 

1. Add solution of chloroplatinic acid (H2PtCl6) which causes 
the precipitation of potassium in the form of the double chloride 
of potassium and platinum (potassium chloroplatinate, KjPtCle). 

2. Add de Konink's reagent^ which is a solution of sodium 
cobaltic nitrite, 6NaN02.Co2(N02)6. This reagent produces a 
yellow precipitate of potassium cobaltic nitrite, 6KN02. 
Co2(N02)6 + XH2O, in a solution containing a potassium salt. 
To hasten the reaction, add a few drops of acetic acid. 

3. Test for sodium in a neutral solution by adding a few drops 
of freshly prepared acid potassium pyro-antimonate solution, 
K2H2Sb207. At first the solution is turbid but, if stirred, de- 
posits a white crystalline precipitate of sodium pyro-antimon- 
ate, Na2H2Sb207. 

Vitali's procedure in testing for caustic alkalies consists in 
shaking the alcoholic extract of the material, prepared as far as 
possible with exclusion of air (see above), with freshly precipi- 
tated and well-washed mercurous chloride. Free alkali black- 
ens this compound. The solubility of mercurous oxide (Hg20), 
the black compound formed, in dilute nitric acid distinguishes 
it from mercuric sulphide. 

Quantitative Estimation of Hydroxides and Carbonates of 
Alkalies. — To determine both the free caustic alkali and that 

^ Prepare sodium cobaltic nitrite by dissolving 10 grams of pure sodium nitrite 
and 4 grams of cobaltous nitrate separately in sufficient water. Mix the solutions, 
add 2 cc. of acetic acid and dilute to 100 cc. with water. 



POISONS NOT IN THE THREE MAIN GROUPS 



187 



I 



converted into carbonate, first determine total alkalinity by 
titrating a portion of the distillation residue -with normal or 
n- hydrochloric acid, using methyl orange as indicator. 

I Then precipitate carbonate in a second portion of the distilla- 
tion residue with barium chloride solution and determine free 
caustic alkali in the filtrate. If the examination shows only 

' alkaline carbonate, this does not exclude the possibility of caus- 
tic alkali having been originally present. 

POTASSIUM CHLORATE 

Toxk Actum. — Large doses (4-10 grams) of potassium cMorate, KClOi, are 
deddedly toxic During the first stage ot intoxication, alteration in the shape of 
the red corpuscles and conversion of oxyhemoglobin in the intact coipuscles into 
brown mc themoglobin take place. Then the red blood corpuscles, at least in a 
case of severe poisoning, change their form, becoming shriveled and undergoing 
decompodtion. Toxicologists (see R. Robert, InloTikationen) ascribe change of 
blood pigment and red blood corpuscles to specific salt action possessed in high 
degree by potassium chlorate. This explanation also accounts for salt diuresis,' 
appearing al the beginning of potassium chlorate poisoning, thereby the blood is 
much thickened. But most notable is the high alkalinity of the urine, resulting 
in decreased alkalinity of the blood plasma. In severe chlorate poisoning so much 
oxybEmoglobtn is changed to melhsemoglobin that the amount of oxygen in 
the blood may drop to i per cent. As a result human beings or animals thus 
poisoned may become asphyxiated from lack of oxygen. Potassium chlorate 
through the action of potassium weakens the heart. In chlorate poisoning the 
blood has a characteristic chocolate -brown color (see above). 

Potassium chlorate taken by the mouth is quite rapidly eliminated by the 
kidneys. After administration of o.t gram of potassium chlorate, chloric add 
appears in the urine in an hour. Most o( the potassium chlorate passes into the 
urine unchanged, only a little of the salt being reduced to potassium chloride. 
During chlorate poisoning, the urine is usually very dark, even black, and may 
contain hxmoglobin and niethxmoglobin. It is frequently opaque and strongly 
alkaline. Upon long standing a dark brown sediment gradually depoats. The 
urine also contains considerable albumin. 

In suspected chlorate poisoning, the urine should if possible receive a thorough 
chemical and microscopical examination. An anuria lasting several days may 
precede death and render an examination of the urine quite impossible. 

Detection of Chloric Add 

To isolate potassium chlorate from organic material, use a 
dialyzer which should be as flat as possible, because the thinn er 
the layer in the inner container and the larger the volume of 

sis = increased secretion of urine. 



188 DETECTION OF POISONS 

water in the outer vessel, the more rapid the diflFusion. Place 
the material to be examined, as parts of organs and stomach 
or intestinal contents, in the inner container of a flat dialyzer 
and pure water in the outer vessel. Allow dialysis to take 
place 5-6 hours without changing the water in the outer vessel. 
Then evaporate the dial)rsate (contents of the outer vessel) to 
dr5aiess in a porcelain dish upon the water-bath. Dissolve the 
residue in a little water and examine the filtered solution for 
chloric acid as follows: 

1. Ihdigo Test — ^Add dilute sulphuric acid and a few drops 
of indigo solution, until there is a distinct blue color. Then 
introduce sulphurous acid drop by drop. If chloric acid is 
present, the blue color changes to yellow or greenish yellow. 
This is a delicate test for chloric acid, given even by o.oi gram 
of potassium chlorate. 

2. Silver Nitrate Test — Add silver nitrate solution in excess. 
If there is a precipitate (AgCl), filter and add a few drops of 
sulphurous acid to the clear filtrate. A chlorate will cause the 
precipitation of more silver chloride. Silver chloride differs 
from silver sulphite in being insoluble in hot dilute nitric acid. 
Sulphurous acid reduces silver chlorate to chloride: 

AgClO, -h 3H,S0, = AgCl 4- 3H,S04. 

3. Free Chlorine Test — A solution containing a chlorate, 
heated with concentrated hydrochloric acid, gives free chlorine. 
The gas passed into potassium iodide solution liberates iodine. 
Shake the solution with chloroform which dissolves iodine with 
a violet color. This test indicates chloric acid only in the ab- 
sence of substances like chromic acid and dichromates which 
also give chlorine with hydrochloric acid. 

If the material is a powder, dissolve in water and filter if 
necessary. A direct test for chloric acid is usually possible with 
such a solution. 

Quantitative Estimation of Chloric Add 

To estimate potassium chlorate quantitatively in urine, dialy- 
sates and other liquids, reduce with zinc dust, or employ 
Scholtz's method. 



POISONS NOT IN THE THREE MAIN GROUPS 189 

1. Zinc Dust Method. — Divide the solution into two equal 
parts. Determine chloride gravimetrically in one portion by 
precipitating and weighing AgCl, or volumetrically by titrating 
according to Volhard's method. 

Determine chloride and chlorate together in the second 
portion. Add 5-10 grams of zinc dust and a little dilute sul- 
phuric or acetic acid, and heat the mixture 0.5-1 hour upon a 
boiling water-bath. Filter and wash the residue with boiling 
water. Acidify the filtrate with nitric acid and precipitate 
chloride with silver nitrate. More chlorine appears in the sec- 
ond than in the first determination. Calculate the percentage 
of potassium chlorate from the difference between the two 
chlorine determinations. One molecule of KClOa upon re- 
duction yields i molecule of KCl and therefore i atom of chlo- 
rine. 

Zinc dust in presence of sulphuric or acetic acid reduces potassium chlorate to 
chloride: 

(a) KCIO, + aZn = KCl + 3ZnO, 

(/3) ZnO + 2CH,.C00H = H,0 + Zn(CH,.COO),. 

2. Method of M. Scholtz.' — This method makes use of the 
reducing action of nitrous acid upon chloric acid: 

HCIO, + 3HNO, = HCl + 3HNO,. 

Add to the solution 10 cc. of nitric acid (sp. gr. 1.2 = 32 per 
cent.) and 10 cc. of 10 per cent, sodium nitrite solution. Let 
the mixture stand for 15 minutes at room temperature. Then 
add 30-50 cc. of o.i n-silver nitrate solution and 5 cc. of satu- 
rated iron alum solution, (H4N)2S04.Fe2(S04)3.24H20. Ti- 
trate excess of silver with o.i n-ammonium sulphocyanate 
solution. 1000 cc. of 0.1 n-AgNOs = 0.1 KCIO3 gram = 
12.245 grams of KCIO3. 

The slight excess of nitrous add has no effect upon the deli- 
cacy of the reaction. Liquids like dialysates of stomach con- 
tents and organs always contain chloride. In that case first 

* Archiv der Pharmazie 243, 353 (1905). 



»l 



190 DETECTION OP POISONS 

determine the amount of chloride in another portion by Vol- 
hard's method. 

H. Hildebrandt^ has adapted Scholtz's method to the ex- 
amination of urine. First completely precipitate chloride in a 
measured volume of urine with silver nitrate in presence of 
nitric acid. Add sodiimi nitrite solution to the clear, chloride- 
free filtrate, as well as more silver nitrate solution, until there 
is no longer a precipitate. Determine as usual the weight of 
silver chloride obtained. 

In the case of urine a larger quantity of nitrous add is decomposed by the urea: 
CO(NH,), + 2HN0j = CO, + 2N, + 2H,0. 
Consequently do not use too little sodium nitrite. 

Behavior of Potassium Chlorate in Putrefaction 

C. BischofiF states that potassium chlorate, mixed with moist, 
organic substances, especially blood, is very soon reduced to 
chloride! BischofiF describes several cases, in which poisoning 
by potassium chlorate had undoubtedly occurred, and yet 
chloric acid could not be detected chemically in parts of the 
cadaver. 

In an experiment, loo grams of blood, 0.5 gram of potassium 
chlorate and 100 grams of water were allowed to stand for $ 
days at room temperatures. Not a trace of chloric acid could 
be detected in the dialysate. BischofiF concludes from this 
experiment that potassium chlorate, mixed with moist organic 
substances, especially with blood, is soon reduced. Conse- 
quently, chloric acid may not be detected, even in cases of 
rapidly fatal poisoning by potassimn chlorate. 

Detection of Chlorate in Meat 

The German law of June 3, 1900, relating to the inspection of beef-cattle and 
meat, forbids the use of chlorates in preserving meat, sausage and fat. The 
official directions prescribed for the chemical examination of meat and fats are aa 
follows: 

Let 30 grams of finely divided meat stand i hour in the cold with 100 cc. of 
water and then heat to boiling. Filter when cold and add sDver nitrate solution 

* Vierteljahrsschrift fllr gerichtliche Medizin 32, 81 (1906). 



I 



POISONS NOT IN THE THREE MAIN GROUPS 191 

bi excess to the filtrate. Add : cc. of lo per cent, sodium sulphite solution and 
3 cc. of concentruted nitric add to 50 cc, of the clear filtrate from the silver pre- 
cipitate and then heat to boiling. If there is a precipitate, insoluble in more hot 
water and consisting of silver chloride, chlorate is present. 

SAHTONIN, StJLPHONAL AMD TRIONAL 

These substances do not find a place in the Stas-Otto process 
on account of their behavior toward cold tartaric acid solutiou 
and ether. Use the following method for their detection. 

Extract the material, neutral or faintly acid with tartaric 
acid, under a reflux condenser with boiling absolute alcohol. 
Filter hot and evaporate the filtrate to dryness upon the water- 
bath. Dissolve the residue in hot water. If the solution is 
colored, digest for some time upon the water-bath with bone- 
black and stir frequently. Filter the hot solution. All of the 
above substances, if present in considerable quantity, crystallize 
in part as the solution cools. Extract the filtrate and any 
crystals thorouglily with chloroform several times. Pass the 
chloroform extract through a dry filter. The residue from 
diloroform may contain santonin, sulphonal and trional, as 
well as acetanilide and phenacetine. 

The chloroform residue may also contain those substances 
extracted in the Stas-Otto process from the acid solution by 
ether. Chloroform completely extracts substances like anti- 
■pyrine, caffeine, acetanilide, phenacetine and salicylic acid. 
As a rule they are purer from this solvent than from ether. 

le chloroform residue may also contain the weak base narco- 
tine. 

SAHTONIN 

Santonin, CjiHuOi. crystallizes in colorless, inodorous, shining leaflets which 
e bitter and melt at 170°. Santonin dissolves in 5000 parts of cold and 150 
parts of boiling water; in 44 parts of alcohol; and in 4 parts of chloroform. All 
these solutions are neutral. It is slightly soluble in ether (r rijo). Light turns 
these crystals yellow. Upon evaporation, an alcoholic solution of the yellow 
modification deposits white santonin. 

Constitution. — Santonin is the internal anhydride (lactone) of aanlonic acid, 
CiiHhO|. Caustic alkalies, as well as caJciurn and barium hydro^tides, dissolve 
n forming salts of this acid. In this case, as with all lactones, the lactone 
s broken as follows: 



192 DETECTION OF POISONS 

CHi CHt 

I H, T H, 

c c c c 

H,C C CH — O. H H,C C CH— OH 

I I I >c6T0K- I I I 

DC C CH— CH'^ DC C C 



H— CH— COOK 

C C CH, C C CH, 

I H, 
CH, 

Santonin Potassium santonate 



CH, 



A solution of a santonate, acidified with hydrochloric add, first gives free 
santonic acid. To isolate this compound from the mixture, extract at once with 
ether. Otherwise, the acid loses i molecule of water upon standing and passes 
into its internal anhydride, santonin. 

Santonin is also a ketone. As such it forms a hydrazone, dsHigOs » N- 
NH.C«H,, with phenylhydrazine and an oxime, Ci,Hi80i = N0H, with hy- 
droxy lamine. 

According to the structural formula above, santonin is a derivative of hexa- 
hydro-dimethyl-naphthalene. Fused with potassium hydroxide, santonic 
acid gives hydrogen, propionic acid and a naphthalene derivative, namely, 
dimethyl-/3-naphthol. 

Behavior in the Organism. — Santonin seems to be incompletely absorbed in the 
body. M. Jaff^^ has administered quite large quantities of santonin to dogs and 
rabbits. He obtained a new substance, called o-oxysantonin (CuHigOJ, from 
the urine of the dog, amounting to 5 or 6 per cent, of the santonin administered. 
He extracted with chloroform considerable quantities of unaltered santonin from 
the feces of the dog. Rabbits can usually tolerate being fed with santonin for 
weeks, and a-oxysantonin is formed only in very small quantity. In the ether 
extract of the rabbit's urine, Jaff6 found a second santonin derivative, /3-oxy- 
santonin, isomeric with a-oxysantonin, with considerable unaltered santonin. In 
these experiments only about half the santonin administered was absorbed by the 
rabbit. 

After administration of santonin, a red pigment called santonin red appears in 
human urine. Even after medicinal doses santonin urine is red, or becomes at 
least scarlet-red to purple on addition of potassium or sodium hydroxide solution. 
Urine containing santonin also becomes carmine red on addition of calcium hy- 
droxide solution. 

Detection of Santonin 

Ether, benzene, or better chloroform, extract santonin only 
from acid solutions. The organic solvent fails to remove this 
compound from an alkaline solution, as it is then in the form 
of a santonate. Santonin is not an alkaloid and forms no pre- 

^Zeitschrift fiir physiologische Chemie 22, 537 (1896-1897). 



POISONS NOT IN THE THREE MAIN CROUPS 



193 



' dpitates with the general alkaioidal reagents, but it gives 
several more or less characteristic color reactions. 

1. Alcoholic Potassium Hydroxide Test — Pure santonin, 
heated with an alcoholic solution of potassium hydroxide, gives 
a fine carmine red color, which gradually changes to reddish 
yellow and finally fades entirely. In this test yellow santonin 
dissolves at once with a yellowish red color. 

2. Sulphuric Acid-Ferric Chloride Test.^ — Heat santonin with 
concentrated sulphuric acid and add a drop of ferric chloride 
solution. The mixture becomes violet. Use about i cc. of 
sulphuric acid to o.oi gram of santonin. 

3. Furfurol-Sulphuric Acid Test^Mix 2-3 drops of alcoholic 
santonin solution with i-z drops of 2 per cent, alcoholic furfurol 
solution and 2 cc. of pure concentrated sulphuric acid. Warm 
this mixture in a small porcelain dish upon the water-bath. 
A purple-red color appears and changes with continued heating 
to crimson-red, blue-violet and finally to dark blue (Thater'), 

^^ Only a few alkaloids and glucosides give distinct color reacUons with furfurol 
^^hnd sulpburii; acid. Substances behaving similarly are veratrine, picrotoxin 
^BEviolet) and piperine (green to blue-gieen, finally indigo-blue). The colors given 
^^w a- and 0-naphlhol with futfurul and sulphurii^ acid arc also characteristic. 



SULPHOMAL 



SulpboDal, CiHtaOtSi. crystallizes in colorless, inodorous and tasteless prisms, 
melting at 135-136° and distilling with slight decomposition at 300*. It is soluble 
Qjj in soo parts of cold and 13 parts of boiling 



1 



parts of ether; and in 6$ parts oE cold and 1 parts of 
boiling alcohol. Sulphonal is freely soluble in chloro- 
form. Especially characteristic of this compound are 
the ease with which it crystallizes and its great stability 
presence of chemical reagents, I'he halogens, halogen adds, alkaline hy- 
ides and carbonates, concentrated sidphuric and nitric adds are without 
the cold. 



^CHr-C— SO,.C,Hi 

H SO,.C,Ki 

Kipr 



Preparation. — The condensation of ethyl mercaptan (2 
molecules) with acetone (i molecule) by means of dry hydrogen 
chloride gas, or concentrated sulphuric acid, results in the forma- 
ion of the ethyl-mercaptole of acetone. The latter compound, 
rith a saturated solution of potassium permanganate 



■ Archiv der Pharmazie 335, i 



USqt). 



194 DETECTION OP POISONS 

in presence of dilute sulphuric* acid, undergoes oxidation with 
formation of sulphonal:^ 



H,Cv HSCH* H,Cv /SCtH, + O, H,C\ /SO,C,Hi 

>C=0+ =H,0+ >C< ► >C< 

H,C/ HSCH* H,C/ \SC,H, + O, HiC^ ^SO,C,H, 



>C=0+ =H,0+ >C< ► >C< 

Ethyl Bthyl-merca] 

mercaptan of acetone 

Detection of Sulj^ioiial 



Acetone Ethyl Bthyl-mercaptole Sulphonal 



Ether, or better chloroform, extracts sulphonal from acid, 
neutral and alkaline solutions. Test the residue left upon 
evaporating these solutions as follows: 

1. Melting-point Test. — Determine the melting point (125- 
126°) of the perfectly pure substance. Crystallization from 
boiling water with the use of a little bone-black easily gives a 
pure product. A mixture of these crystals with pure sul- 
phonal should also melt at 125-126°. 

2. Reduction Test — Sulphonal heated in a test-tube with 
powdered wood charcoal gives the characteristic odor of ethyl 
mercaptan. 

3. Detection of Sulphur. — (a) With Sodium. Fusion of 
sulphonal in a dry test-tube with a little metallic sodium pro- 
duces sodium sulphide. Dissolve cautiously (unaltered metallic 
sodium!) the cold melt in a little water, filter and test the 
filtrate with sodium nitroprusside solution for sulphide (see 
page 23). 

(b) With Potassium Cyanide. — Fuse i part of sulphonal 
and about 2 parts of pure potassium cyanide in a dry test- 
tube. Note the penetrating odor of ethyl mercaptan (C2H6.SH) . 
Potassium sulphocyanate is also a product of the reaction. An 
aqueous solution of the melt, acidified with dilute hydrochloric 
acid, becomes deep red with 1-2 drops of ferric chloride solution. 

(c) With Powdered Iron. — Sulphonal heated with pure pow- 
dered iron free from sulphur gives a garlic-like odor. Add 

* Sulphur in the su]phone group = SO2 is most likely sexivalent, corresponding 
to the atomic grouping I, and not quadrivalent, as in II: 



VI ^O IV yO 

I. =85 



\ ; n. =s<;i 




POISONS NOT IN THE THREE MAIN GROUPS 195 

lydrochloric acid to the residue. Hydrogen sulphide evolved 
ikens lead acetate paper. 

Detectton of Sulphonal in Urine 

SulphoD^ is cumulative in its action. Therefore continuous administratioD 
long time of large doses may result in the collection of a considerable 
quantity of this compound in the organism. Most of the sulphonn] taken ap- 
pears in the urine as ethyl-sulphonic acid, CiHi-SO,OH.' The formation of 
this acid causes an increase of ammonia in the urine during sulphonal intoxi cation, 
as docs administration of mineral acids. 

Sulphonal occurs in urine in detectable quantity only following considerable 
doses, especially when they have been taten without interruption. Such urine 
is often dark red to gamct-brown from hsmaloporphyrin. But thi> decom- 
podtion product of blood pigment appear? in urine only succeeding severe 
poisoning by sulphonal, and even then its occurrence is rare. 

To isolate sulphonal from urine, evaporate looo cc. to one-tenth its volume, 
several times with large quantities of elher. Pass the ether extracts, 
after they have settled In a dry flask for several hours, through a dry hlter and 
Evaporate the residue with 30 -30 cc. of 10 per cent, sodium hydroiide 

lution to dryness upon the water-bath. This will remove coloriug matter, 
extracted from urine by ether, but will not affect the sulphonal. Extract sul- 
phonal from the alkaline re^due with ether. Evaporate the solvent, and sul- 
phonal will remain pure and almost colorless. Determine the melting point of 
this residue, and make the other tests for sulphonal. 



■ after 
Hi>ti]. 

^Boiuu 

HoEtrai 



I 



Detection of Heematoporphyrin in Urine 

Coloring matters have been observed in red, brownish red to cherry-red 
ines, which quite probably are identical with hEmatoporphyrin. The 
spectroscopic examination of such urine is made in the following manner. Add 
sodium hydroiide solution, drop by drop, to about 500 cc. of urine, until the 
reaction is strongly alkaline, and then add a little barium chloride solution. 
Filter after a while, and wash the precipitate well. Extract the predpitate 
upon the filter with hot alcohol, containing a few drops of dilute sulphuric add, 
.\ spectroscopic examination of this filtrate can be made directly with a Brown- 
ing pocket spectroscope. Add bematoporphyrin solutions are violet; when 
more concentrated, they have a cherry-red color, and show the characteristic 
spectrum with two absorption bands (see page 199). If the acid, alcoholic 
solution is saturated with a few drops of ammonium or sodium hydroxide 
solution, the spectrum of alkaline hematoporphyrin solution with its four ab- 
sorption bands appears. Traces of hEmatoporphyrin very frequently appear 



4 



It should not be confused with ethyl-sulphur 



196 



DETECTION OF POISONS 



in normal urine. It has been observed more abundantly, at times, in urine 
during chronic sulphonal poisoning. 

TRIONAL 

Trional crystallizes in colorless, shining leaflets melting at 76^. It is soluble 
in 320 parts of cold, but more easily soluble in hot water. It is also soluble in 

alcohol, ether and chloroform. The aqueous solution is 

C^i\ y^tX:^k neutral and bitter. In the latter respect it differs from 

CjHj/ ^SOa C2H§ sulphonal which is tasteless. Trional gives the sulphonal 

reactions. Trional is completely decomposed in the 
organism and the danger of cumulative action is much less than in the case of 
sulphonal. Moreover, haematoporphyrin has almost never been observed, even 
following considerable doses of trional and after uninterrupted use for weeks. 



Active Organic Substances^ Rarely Occurring in Tozicological 

Analysis 

CANTHARIDIN 

Cantharidin, CioHi204> is the active vesicating principle of Spanish fly 
(Lytta vesicatoria) and is present to the extent of 0.8-1 per cent. Cantharidin 

forms colorless, shining, neutral, rhombic leaflets, 
melting at 218** and subliming at higher temperature 
in white needles. It is almost insoluble even in 
boiling water. Adds, as tartaric add, increase its 
solubility in water, though cantharidin is not a base. 
It dissolves with difficulty in cold alcohol (0.03: 100 at 
18°) and in ether (o.ii : 100). Chloroform (1.52 : 100), 
acetone and acetic ether are its best solvents. It is 
as good as insoluble in petroleum benzine. 
Constitution. — According to H. Meyer' cantharidin has the structural formula 
shown above. This compound is at the same time a monobasic add and a 
/3-lactone. Potassium or sodium hydroxide breaks the labile /3-lactone ring. 
Cantharidin passes into solution as the alkali salt of diabasic cantharidic add, 
CioHi40i: 

H K 

C CH,— COO H 



H 

C CHj.COOH 

H2C c— o 

CH2 

\l 

C— CO 



H2C 



\ / 

C 

H2 



HjC C— 




CH2 






\ ' 




H2C C— CO 


\ / 


c 


H2 




Cantharidin 





+ 



OH 


H 




H 


C 




/ 


\ /CH2— COOK 




HjC 




CH2 


^OH 


H 


\ 




OK 


H2C C— COOK 




\ / 




c 




H2 




Po 


tassit 


urn cantharidate 



+ H,0. 



* The toxic substances considered in this place have been arranged in alpha 
betical order. 

* Monatshefte ftir Chemie 18, 393 (1897) and 19, 707 (1898). 



POISONS NOT IN THE THREE MAIN GROUPS 197 

Potassium cantharidate, CioHuOtKi.aHiG, recently recommended for 
phthisis, and sodium cantharidate, CioHisOiNai.2HtO, are well crystallized 
salts. Mineral add first sets cantharidic acid free from these salts. The latter 
soon loses a molecule of water, passing into its internal anhydride, cantharidin. 



i/i 



H H 

C CHr-COOH C CH,— COOH 

\ / / \ / 

C— OH HaC C— O 

CHj 



+ H,0 



I \l l... \ ^ 

H,C C— COOH HtC C— CO 

\ / \ / 

c c 

H, H, 

Cantharidic acid Canthandin 

Cantharidin, heated with hydriodic acid at 100**, or treated at room tem- 
perature with chlorosulphonic acid, Cl-SOj-OH, changes into the isomeric 
cantharic acid, C10H11O4, crystallizing in colorless needles melting at 275^ 
This acid is not a vesicant. Heated for 3 hours at 135** in sealed tube with 
acetyl chloride, cantharic acid jdelds another isomer of cantharidin called iso- 
cantharidin (Anderlini and Ghiro).^ The latter crystallizes from alcohol in 
large colorless leaflets melting at 76**. There is a close relationship between o- 
xylene and cantharidin, for the latter, heated at 400° with calcium hydroxide, 
gives a dihydro-o-xylene, CgHn, called cantharene, and also o-xylene, C«Hr 
(CHs)i, and xylic acid. FinaUy, cantharidin, heated with an excess of phos- 
phorus pentasulphide and distilled, gives pure o-xylene. (J. Piccard.)* 



Detection of Cantharidin 

Evaporate a liquid, or material containing much moisture 
(organs, stomach or intestinal contents, etc.), to dryness upon 
the water-bath. Dragendorflf directs repeated extraction of the 
finely divided material with alcohol containing sulphuric acid. 
Filter the extracts, add one-sixth their volume of water and 
distil the alcohol. Extract the residue 2-3 times with chloro- 
form and shake the chloroform extracts with water to remove 
adherent acid. Finally separate the chloroform from water, 
distil and examine the residue for cantharidin. Since this com- 
pound gives no characteristic chemical reactions, employ the 
physiological test for identification. Dissolve the chloroform 
residue, unless fatty substances are present, in a few drops of 

* Berichte der Deutschen chemischen Gesellschaft 24, 1998 (1891). 
«Ibid., 12,577 (1879). 



198 DETECTION OF POISONS 

hot almond oil. Bind a cloth, saturated with this solution, 
upon the upper arm or breast by means of adhesive plaster. 
Cantharidin reddens the skin and sometimes raises blisters. 
Even 0.14 mg. of cantharidin causes blistering. Salts of can- 
tharidic acid also have a vesicating action. 

To detect cantharidin in blood, brain, liver and other material 
rich in protein, E. Schmidt boils with dilute potassiimi hydroxide 
solution (i gram of KOH in 15 cc. of water), until the mass is 
homogeneous, acidifies with dilute sulphuric acid and extracts 
thoroughly with hot alcohol. The procedure in other respects 
is as described above. 

Cantharidin is said to resist putrefaction. 

CYTISINE 

Cytisine, C11HUN2O, occurs in the ripe seeds of Golden chain (Cytisus Labur- 
num) which contain about 1.5 per cent. Cytisine and the alkaloid originally 
called ulexine, isolated from the seeds of Ulex europaeus, are identical (A. 
Partheil). 

Preparation. — Extract powdered ripe laburnum seeds with 60 per cent, alcohol 
containing acetic add. Distil the alcohol from the extracts, pour the residue 
through a moist filter and precipitate extractive and tannin substances with 
lead acetate solution. Filter, add potassium hydroxide solution to the clear 
filtrate and extract cytisine with chloroform. Distil the chloroform which 
usually deposits cytisine as a radiating crystalline mass. If purification is neces- 
sary, recrystallize the residue from absolute alcohol or boiling ligrOin. Sub- 
limation in a partial vacuum also purifies crude cytisine. 

Cytisine crystallizes in large, colorless, tasteless prisms, melting at 152—153** 
and subliming at a higher temperature, if carefully heated. It dissolves freely 
in water, alcohol, chloroform and acetic ether; less easily in commercial ether, 
benzene and acetone; and is insoluble in petroleum ether and absolute ether. 

Cytisine is a strong secondary base and very toxic. Although capable of com- 
bining with I or 2 molecules of hydrochloric acid, this compound behaves in 
other respects like a monacid base. Only the salts containing one equivalent 
of acid crystallize weU. Nitrous acid converts this secondary base into nitroso- 
cytisine, CnHijON-NO, which crystallizes in needles. Nitrous fumes appear, 
if cytisine is warmed upon the water-bath with twice the amount of concentrated 
nitric acid, and the solution at once becomes reddish yellow to brown. This 
solution poured into water gives a precipitate of nitro-nitroso-cytisine, CnHnON- 
(N02)N-N0. This compound crystallizes from water in pale yellow scales 
melting at 242-244°. 

Toxic Action. — Cytisine produces convulsions, its action in this respect being 
very similar to that of strychnine. But unlike the latter alkaloid it also irri- 
tates the gastro-intestinal mucosa even causing bloody inflammation. Cytisine 



POISONS NOT m THE THREE litAlN GROUPS 



:99 



slso differs from strychnine in stimulating the vomiting center. Consequently, 
after doses of cytisine or laburnum preparations, human bdngs and animals 
capable of emeais thus rid the organism of a large part of the poison. Like 
strychnine cytisine stimulates the respiratory and vaso-motor centers. Finally 

»H in strychnine intoxication death results from paralysis of these two centers 
A part of the cytisine leaves the organism unchanged and appears in the urine 
(R. Kobert.) 



Detection of CTtisine 



Prepare an aqueous tartaric add solution of stomach con- 
tents, vomitus or parts of organs, following the general pro- 
cedure for alkaloids, To remove final traces of fatty acids and 
fat, shake this solution well with ether. Withdraw the aqueous 
ijwlution, make alkaline with sodium hydroxide solution and 
:ract thoroughly with chloroform or isobutyl alcohol. Evap- 
orate the chloroform or isobutyl alcohol extracts and test the 
residue as follows for cytisine: 

1. Van der Moer's' Test — Ferric chloride solution colors 
cytisine and its salts blood red. Dilution with water, or acidi- 
fication, discharges this color. Hydrogen peroxide also pro- 
duces the same result. The solution containing hydrogen 
peroxide, warmed upon the water-bath, becomes intensely 

ilue. 

2. A. Ramverda's^ Test^ — A little nitrobenzene, containing 
dinitro-thiophene, poured upon cytisine gives a fairly stable, 
brilliant red- violet color. 

A similar color given by coniine is very unstable. 

3. Nitro-Tfitroso-Cytisine Test— Nitro-nitroso-cytisine (see 
ibove), formed by concentrated nitric acid, serves to detect 

ail quantities of this alkaloid. Nitro-nitroso-cytisine dis- 
ilves with difficulty in 94 per cent, alcohol and crystallizes 
■om this solvent in microscopic prisms. Flat, tabular crystals 
form from 50 per cent, alcohol which is a better solvent. The 
solubility of nitro-nitroso-cytisiae in concentrated hydrochloric 
acid indicates basic properties, but they are feeble, for dilution 
with water precipitates this compound unchanged. 

■ Berichte der Deutschen pharniaieutischeo Gesellschaft, s, 267 (1895). 
' Chemisches Zentral-Blatt, i goo, II, j6a. 



tat, 
■ ora 






200 DETECTION OF POISONS 

THE DIGITALIS GLUCOSIDBS 

The digitalis plant (Digitalis purpurea L.) contains in all its 
parts, but especially in the leaves and seeds, medicinally useful 
substances belonging to the glucoside group. Thus far three 
digitalis glucosides have been isolated as well characterized, 
crystalline compounds of homogeneous composition. These are 
digitalin in a narrower sense (= Digitalinum verum crjrstal- 
lisatimi Kiliani) CsfiH^eOn; digitoxin, Cs4HmOii; and digitonin, 
C(6H94028 or C(4H92028- A fourth glucoside called digitalein 
seems not to have been obtained wholly pure as yet. 

Digitonin, C&6H94O28 or CsiJtl9202B,^ occurs almost exclusively 
in digitalis seeds, the leaves containing at most only traces. 
Digitonin, classified at present with the saponins (see page 213), 
crystallizes from alcohol in fine needles soluble in 50 parts of 
50 per cent, alcohol. Even very dilute hydrochloric acid 
hydrolyzes digitonin into digitogenin, dextrose and galactose:* 

CMH94OM + 2H2O = CiHmO. -f 2C.Hi,0. + 2C,Hi,0,(?). 

Digitonin Digitogenin Dextrose Galactose 

Digitonin crystallizes from alcohol in fine needles which soften 
at 235° and become yellow. Digitonin is not a cardiac poison. 
Pure digitonin and concentrated sulphuric acid, upon addition 
of a little bromine water, give a color which becomes intensely 
red. 

Digitozin, CsiHsiOn, occurs almost exclusively in digitalis 
leaves. This very active and highly toxic compound is almost 
wholly insoluble in water and ether but soluble in alcohol and 
chloroform. Consequently ether precipitates it from chloro- 
form solution. Digitoxin crystallizes from 85 per cent, alcohol 
in leaflets melting at 145°. Alcoholic hydrochloric add hy- 
drolyzes it forming digitoxigenin and digitoxose: 

C,4H»40ii + H,0 = C«H„04 + 2C.Hn04. 

Digitoxin Digitoxigenin Digitoxose 

Digitoxin dissolves in concentrated sulphuric acid with a 
brownish or greenish brown color unchanged by bromine. 

* The results obtained by A. Windhaus (Berichte der Deutschen chemischen 
Geselbchaft 42, 238 (1909) favor the formula C»kH«40i8 for digitonin. 

* H. Kiliani, Berichte der Deutschen chemischen Gesellschaft 24, 340 (1891). 



POISONS NOT IN THE THREE MAIN GROUPS 201 

Kiliani's Digitozin Test^ — Dissolve a trax:e of digitoxin in 
3-4 cc. of glacial acetic acid containing iron (loo cc. of glacial 
acetic acid and i cc. of 5 per cent, ferric sulphate solution). 
Cautiously add sulphuric acid containing iron (100 cc. of sul- 
phuric add and i cc. of 5 per cent, ferric sulphate solution) in 
about the same quantity as an under layer. A dark zone ap- 
pears where the two solutions meet, above which after a few 
minutes a blue band is visible. After some time the entire 
acetic acid layer becomes deep indigo -blue. 

Digifalin, CssHseOu, according to Kiliani occurs only in 
digitalis seeds. It is soluble in water (i : 1000) and very active. 
Boiling with very dilute hydrochloric acid hydrolyzes its alco- 
holic solution into digitaligenin and two sugars, namely, dex- 
trose and digitalose:* 

CuHmOu -f H,0 = C«H,oO, -f C,Hi,0, + 2C7H14O, 
Digitaiin Digitaligenin Dextrose Digitalose 

Test for digitaiin as follows : 

1. Concentrated sulphuric add colors pure digitaiin orange 
yellow. This solution soon becomes blood red, changing upon 
addition of a little bromine water to cherry and blue-red. A 
drop of nitric add or ferric chloride solution will do as well as 
bromine water. This test after 1-2 hours is surer and more 
permanent, if a trace of digit alin is dissolved direct in concen- 
trated sulphuric add and nothing else is added. 

2. Concentrated hydrochloric add dissolves digitaiin with a 
golden yellow color, changing with heat to garnet or violet-red. 

At present nothing definite is known regarding the fate of digitalis glucosides 
in the human organism, or the products into which they are changed or the 
forms in which they are eliminated. In the case of human beings elimination 
of the three active substances has never been observed. Moreover, R. Robert 
has not been able to detect anything active in the urine of animals except in 
isolated cases. Thus far it has not been possible to find any of the digitalis 
compounds mentioned above in blood or animal organs. In a toxicological 
analysis especial attention would have to be given to vomitus and the contents 
of the gastro-intestinal tract. But there is slight chance of detecting the digitalis 
bodies in such material. 

* Archiv der Pharmazie 234, 273-277 (1896). 

' H. Kiliani, Berichte der Deutschen chemischen Gesellschaft 31, 2454 (1808). 



DETECTION OF POISONS 



I 



ERGOT 

OfScinal ergot (Secale cornutum) is the sderotium {compact myccliuin or 
spawn) of CUiviceps purpurea coUecled from r>'e shortJy before the fruiting period 
and dried at gentle beat. Ergot is commonly known as an abortifacicnt and in- 
torications have occurred ftom its use. Consequently examinations for legal pur- 
poses may require its detection in powders and other mijtures. Our knowledge 
of the constituents of ergot is still very defective notwithstanding several ex- 
haustive investigauons. Ergot alkaloids, as ergotine, eigotinine, comutine,picro- 
Gclerotine, were described long ago. But, with the possible exception of ergotinine 
(Tanret, C. C. Keller), the preparations were not entirely pure. Ergot conuina 
in addition to alkaloids other peculiar chemical substances which have received 
but little attention. They have not the characteristic physiological action of 
ergot but, like the gngment sdererythrin, are useful for purposes of identification. 
Among these substances belong sphacelic acid and sderoUc add, according to R. 
Robert a very poisonous resin having add properties. 

Alkaloids. — The most recent' researches upon ergot mention as well character- 
ized bases ergotinine, CitHiiNtOt, and hydro- ergotinine, CjiHiiNtOt. Barget 
calls the latter ergotoxine. Ergotinine crystallizes from alcohol in long needles 
mdting at about «39° when heated rapidly. This compoimd dissolves in ja 
parts of boiling alcohol; in i.i parts of ether; and is readily soluble in chloroform. 
Crystalline salts of this base have not yet been prepared. Hydro-crgotinine 
( — hydrate of ergotinine), obtained as a crystaUine phosphate from ergotinine 
mother Uquors by means of alcohol and phosphoric add, is a white powder soften- 
ing at 155° and melting at 163-164°, Though freely soluble in alcohol, it dis- 
solves but slightly in ether. As a rule the salts of hydro- ergotinine (ergotoxine) 
crystallize well.' By preparing a cold methyl alcohol solution of hydro-ergotinine 
and boiling this solution for several hours under a return condenser, F. Etoft has 
converted this substance completdy into ergotinine. On the other band, ergoti- 
nine in dilute acetic add solution passes back almost entirely into hydro-ergotinine 
within 10 days. As an indication ol ptirity, a solution of hydro-ergotinine in a 
pail^ of cold methyl alcohol after several days standing should not deposit 
crystals (ergotinine) nor become green. Solutions of both alkaloids are fluores- 
cent. According to Keller the play of colors with sulphurc acid and ferric chloride 
is characteristic of ergotinine (see below). 

Physiological Action of the Alkalotde. — Ergotinine and hydro-ergotinine accord- 
ing to A. Jaquet produce convulsions and gangrene. They are not, however, the 
cause of the spedfic uterine contraction characteristic of ergot. Keller's cornutine 
according to Kraft is identical with ergotinine, according to G, Barger and H. H. 
Dale' with ergotinine, which is impute from ergotoxine (hydro-ergotinine). 
The English investigators beheve that the physiological effects observed with 
ergotinine ore due to adhering ergotoxine. The latter is readily formed when the 
difficultly soluble ergotinine is brought into solution by means of gladal acetic acid, 
phosphoric acid, or a little sodium hydroxide solution. Ergotoxine according to 

' F. Kraft, Archiv dcr Pharmazie 14^,336 (1906) and G. Barger, JounuJ of the 
Chemical Sodety gi, 337. 

' G. Barger and F. H. Carr, Proceedings of the Chemical Sodety »3, »7 

' Bio-Chemical Journal 3, 340. 



POISONS NOT IN THE THHEE MAIN GROUPS 203 

Bftrgec and Dale producM the cEFecU typical of ei^ot, causing powerfu) contiac- 
lion of the uterus and later abortion. 

SclererythriD. — This ig the pigmeni of the outer coat of ergot. E. Schmidt 
gives the foliowing directions for its isolation. Extract freshly powdered ergot 
with ether to remove fat. Then moisten the powder with water containing tar- 
taric add, dry and exttact Rilb 95 per cent, alcohol. Filter and distil the alcohol. 
Extract the residue with ether. This solvent now dissolves sdererytlirin which 
can be precipitated by means of petroleum ether. 

Sdereryttuin is an amorphous red powder which can be sublimed. It is in- 
soluble in water but soluble in absolute alcohol and glacial acetic add. This 
lubsumce behaves like an add, dissolving in caustic alkalies, ammonia, and 
allcaline carbonate and bicarbonate solutions with a red or red-violet color. 
Owing to presenile of sdererythrin, ether, if shaken with powdered ergot moistened 
with tartaric acid solution, becomes red. If such on ether solution of sderery- 
ttuin is shaken with sodium hydroxide solution, the pigment dissolves in the 
latter which then becomes red. Solutions of ihb pigment show characteristic 
absorption bonds in the spectrum. Moreover the pigment gives blue-violet 
predpitates with solutions of caldum hydroxide, barium hydroxide and lead 
acetate. The predpitate with stannous chloride is currant-red; with copper sul- 
phateapure violet; with ferric chloride a deep green; and with chlorineor bromine 
water a lemon- yellow. 

■ Detection of Ergot tn Flour, Bread and Powders 

This examination usually consists in undertaking to detect 
by chemical and physical means the red pigment sclererythrin 
which is characteristic of ergot. The property possessed by 
this substance of passing from ether into a solution of an alkaline 
hydroxide or bicarbonate is especially valuable for purposes of 
identification. 

II. Detection of Sclererythrin. — Shake frequently and let 
10 grams or more of flour stand for a day in a closed flask with 
30 cc. of ether and about 15 drops of dilute sulphuric acid (1:5). 
Then pass the ether through a dry paper, wash the residue with 
alittle ether and shake the filtrate thoroughly with 10-15 drops 
of cold saturated sodium bicarbonate solution. If the flour con- 
tains ergot, the aqueous layer separates with a violet color. 
R. Palm extracts the flour at 30-40° with 10-15 times its 
volume of 40 per cent, alcohol containing a few drops of am- 
monia. E-xpress the liquid, filter and add basic lead acetate 
solution to the filtrate. Press the precipitate between filter 
paper and warm while still moist with a little cold saturated 



204 DETECTION OF POISONS 

borax solution. A red-violet color appears, if the flour con- 
tains ergot. 

2. spectroscopic Examination.— This test gives a positive 
result, if the material (powdered ergot, flour, bread) contains 
more than o.i per cent, of ergot. Examine spectroscopically 
the alkaline and acid solution of the pigment. The red solu- 
tion, prepared in Test i by means of ether containing sulphuric 
acid, shows two absorption bands. One lies in the green be- 
tween E and F but nearer E and a second broader band in the 
blue midway between F and G. Then render the solution 
alkaline with ammonia. Three absorption bands should ap- 
pear. The first lies between D and E, the second at E some- 
what to the right and the third to the left of F. 

3. Choline. — Ergot powder, warmed with dilute potassium 
hydroxide solution, gives the characteristic odor of trimeth- 
ylamine, (CH3)8N,^ due to decomposition of choline in ergot. 

/CH2.CH,.0H 
(CH,),N< 

Occasionally flour that does not contain ergot may give an 
odor when heated with potassium hydroxide solution. 

4. Detection and Quantitative Estimation of Ergotinine (C. C. 
Keller). — Dry finely powdered ergot over lime, place 25 grams 
in a Soxhlet tube and completely extract fat with petroleum 
ether. Dry the powder at gentle heat, add 100 grams of ether 
and after 10 minutes shake well with milk of magnesia ( i gram 
of MgO and 20 cc. of water). Shake repeatedly during an hour 
and then pass 80 grams of the ether solution ( = 20 grams of 
ergot) through a covered folded filter into a separating funnel. 
Shake the ether in succession with 25, 15, 10 and 5 cc. of 0.5 
per cent, hydrochloric acid. Pour the hydrochloric add ex- 
tracts, now containing the ergot alkaloids, through a small 
moistened filter.^ Add ammonia until alkaline and extract 

^ The so-called com smut (Ustilago Maidis), said to cause effects similar to those 
of ergot, also gives the trimethylamine odor when warmed with potassium hydrox- 
ide solution, for it contains appreciable quantities of choline. 

* Clarify the filtrate from these hydrochloric acid extracts, if not dear, by 
agitation with a little talcum powder, previously treated with hydrochloric add 
and thoroughly washed with water. Then filter again. 



POISONS NOT IN THE THREE MAIN GROUPS 205 

the solution twice with about half its volume of ether. Let 
the ether extract settle in a dry flask, then filter into a dry 
weighed flask and wash the filter with a little ether. Distil 
the ether and dry flask and residue at ioo° to constant weight. 
Good German ergot contains 0.13-0.16 per cent, and Russian 
ergot 0.22-0.25 per cent, of the alkaloid. 
To detect ergot alkaloid qualitatively, proceed as follows: 

(a) Dissolve a part of the residue in i cc. of concentrated 
sulphuric acid and add a trace of ferric chloride solution. The 
mixture is orange-red and becomes at once deep red but the 
margin appears bluish to bluish green. 

(b) Dissolve a part of the residue in about 4 cc. of glacial 
acetic acid and add a trace of ferric chloride solution. Cau- 
tiously add this mixture to concentrated sulphuric acid as an 
upper layer. If ergotinine is present, a brilliant violet color 
appears where the two liquids meet. 

OPIUM 
Detection of Meconic Acid and Meconine 

Since it is comparatively easy to procure small quantities of 
opium preparations, especially the tincture, poisoning from this 
source is possible. Consequentiy, it is often desirable to 
recognize the presence of opium itself. Detection of the 
alkaloids narcotine and morphine, always present in opium in 
considerable quantity, affords partial evidence of the presence 
of this substance. Moreover, opium always contains two non- 
basic substances, meconic acid and meconine. Detection of 
these two compounds in conjunction with narcotine and 
morphine definitely determines the presence of opium. 

Meconic Acid, C7H4O7 = C3H02(OH)(COOH)2, is an oxy- 
pyrone-dicarboxylic acid (II) and therefore a derivative of 
pyrone (I): 









c 


c 


/\ 


/\ 


HC CH 


HC C— OH 


I. 1 


II. 


HC CH 


HOOC— C C— COOH 


\/ 


\/ 








Pyrone 


Meconic acid 



206 DETECTION OF POISONS 

Meconic add crystallizes in plates or prisms with 3 mole- 
cules of water. It is easily soluble in hot water and alcohol. 
A solution of a. ferric salt turns a meconic add solution dark red. 

To detect meconic add, extract a portion of the material 
with alcohol containing a few drops of hydrochloric add. Filter 
and evaporate the filtrate upon the water-bath. Dissolve the 
residue in a little water and heat the filtered solution to boil- 
ing with excess of calcined magnesiimi oxide. The solution 
contains magnesium meconate. Filter hot to remove undis- 
solved magnesium oxide, evaporate the filtrate to a small 
volume and addify faintly with dilute hydrochloric acid. Add 
a few drops of ferric chloride solution. A blood-red color 
appears, if meconic add is present. Warming with hydro- 
chloric add does not discharge this red color, in which respect 
it differs from the red color caused by acetic add. This color 
differs from that caused by sulphocyanic add in not being 
affected upon addition of gold chloride. But stannous chloride 
reduces ferric to ferrous oxide and discharges the color. Nitrous 
add, however, at once restores it. 

These tests permit the identification of meconic acid in an 
extract from only 0.05 gram of opium. 

Meconine, C10H10O4. — Opium contains only 0.05-0.08 per 
cent, of this compound. It forms small prisms, melting at 102° 
and subliming at higher temperature without decomposition. 
Meconine dissolves freely in alcohol, ether, benzene and chloro- 
form, but less easily in water. Alkalies convert meconine 
into easily soluble salts of meconinic acid, CioHwOs. This 
monobasic add cannot exist free but changes to meconine when 
liberated from its salts by a mineral acid. Meconine, formed 
by abstracting a molecule of water from meconinic add, is 
therefore the internal anhydride (lactone) of meconinic acid : 

CHr-biH CHr-0 



c , i 

/\ i... ^\ 

HC C— CO iOH HC C— CO 

I II I II 

HC C— OCHs HC C— OCH, 

\/ \/ 

C C 

I I 

OCH, OCH, 

Meconinic acid Meconine 



POISONS NOT IN THE THREE MAIN GROUPS 



207 



To detect meconine, extract the material with alcohol con- 
taining sulphuric add. Filter and evaporate the filtrate to a 
syrup upon the water-bath. Dissolve the residue in water and 
extract meconine from this add solution with benzene. Evapo- 
ration of the solvent frequently gives crystals of meconine. To 
detect meconine, dissolve in a little concentrated sulphuric add. 
The solution is green but turns red within two days. If the 
green sulphuric acid solution, or that which has turned red upon 
standing, is carefully warmed, a fine emerald-green color ap- 
pears, passing through blue, violet and finally back to red. 

Seleniotts-Sulphuric Add Reagent for Opium Alkaloids^ 

Prepare the reagent used in these tests by dissolving 0.5 
gram of selenious add (H2Se03) in 100 grams of pure concen- 
trated sulphuric add. This reagent is espedally delicate for 
opium alkaloids, detecting even traces of morphine and codeine 
(0.05 miUigram), as well as of papaverine (o.i milligram). 
Selenious-sulphuric acid gives the following color reactions 
with the commoner opium alkaloids: 




Morphine 

Apomorphine . . . 
Codeine 

Narceine 

Narcotine 

Papaverine 

Thebaine 



Blue; then permanent blue 
green to ohve green. 

Dark blue violet. 

Blue qUickly changing to 
emerald green and later tol 
permanent olive green. 

Faint greenish yellow; then 
violet. 

Greenish steel blue; later 
cherry-red. 

Greenish, dark steel blue; 
then deep violet. 

Deep orange gradually fad- 
ing. 



Brown. 

Gradually dark brown. 
Steel blue; then brown. 



Dark violet. 
Cherry red. 
Intense dark violet. 
Dark brown. 



^ Mecke, Zeitschrift ftir dffentliche Chemie 5, 350 (1899) and Zeitschrift ftir 
analytische Chemie 39, 468 (1900). 



208 DETECTION OF POISONS 



PAPAVERmB 

Papaverine, CtoHnNOi, constitutes about 0.5 — z per cent of opium. When 

crude it is usually mixed with narcotine. To remove the latter, prepare the acid 

II II oxalate of papaverine which dissolves with diffi- 

C C culty in water. Crystallize this salt from boiling 

^\/\ water until it dissolves in concentrated su^>huric 

HjC.O C C CH add without color. Convert papaverine oxalate 

II Q Q ^ ^ -^ into the hydrochloride by treatment with calcium 

\/\^ chloride and then liberate the alkaloid with am- 

C C monia. This product crystallized from alcohol is 

°- i pure papaverine. 

I * Papaverine crystallizes in colorless prisms mdt- 

C ing at 147°. This alkaloid is insoluble in water; 

^\ soluble with difficulty in ether (1:260), cold 

"-y ^"- alcohol and benzene; but freely soluble in hot 

IIP Q Q Qii alcohol, acetone and chloroform. These solutions 

\/^ are neutral, not bitter, and optically inactive. 

C Papaverine is a weak base which dissolves in but 

I does not neutralize acetic add. Ether partially 

* extracts it from an aqueous tartaric add solution 

and completely extracts it from alkaline solution. Consequently this alkaloid 

appears in the Stas-Otto process in ether extract B. Chloroform extracts 

papaverine with almost as much ease from an add solution as from one that 

is alkaline. 

Constitution. — Papaverine is a monadd, tertiary base which combines with 
alkyl iodides forming crystalline addition products. As it forms no acetyl deriva- 
t ive with acetic anhydride, free hydroxyl is not present. But there are probably 
four methoxyl groups, for it loses four methyl groups when treated with hydriodic 
add according to Zeisd's method. Consequently all the oxygen atoms in papa- 
verine are present as methoxyl groups. The researches of Guido Goldschmiedt, 
extending from 1883 to 1898, have completdy explained the constitution 
of papaverine. Moderate oxidation with potassium permanganate and sidphuric 
add gives papaveraldine, CsoHi gNO^, without breaking the carbon chain. Fusion 
with potassium hydroxide breaks the latter into nitrogen-free veratric add and 
the nitrogenous base dimethoxy-isoquinoline:^ 



^ Isoquinoline (II) is isomeric with quinoline (I) and like the latter is a monadd, 

tertiary base: 

I. H H II. H H 

C C C C 

/\/\ /\/\ 

HC C CH HC C CH 

I II I I II I 

HC C CH HC C N 

\/\^ \/\/ 

C N C C 

H H H 

Ouinoline Isoquinoline 



POISONS NOT IN THE THREE MAIN GROUPS 



209 



H H 
C C 

HtCQ-C C CH 

H,CO— C C N 

YY 

H I H 
00+. OK 

HC CH 
HC C— OCH, 

Y 

OCH, 

Papaveraldin 



H H 
C C 

H,CO— C C CH 

HiCO— C C N 

C C 
H H 

Dimethoxy-iaoquinol ine 
COOK 

I 

c 

/\ 

HC CH 

HC C— OCH, 

Y 



i. 



>CH, 

Veratric acid 



Detection of Papaverine 

The following general reagents precipitate papaverine in a 
dilution of i : 10,000: phospho-molybdic acid, potassium bis- 
muthous iodide and iodo-potassium iodide. 

The following still give precipitates in a dilution of i : 5000: 
tannic acid, gold chloride and potassium mercuric iodide. 

The following special tests should be made: 

1. Concentrated Sulphtiric Acid. — The cold colorless solution 
of papaverine in this acid becomes dark violet upon gentle 
warming. But even a cold solution of impure papaverine in 
this acid is violet. 

2. Froehde's Test — The solution of pure papaverine in this 
reagent is green. Blue, violet and finally a brilliant cherry red 
color appear upon wanning the solution. 

3. Solutions of this alkaloid in concentrated nitric acid, or 
Erdmann's reagent, are dark red. 

Heat to boiling a solution of i part of papaverine with 10 parts of nitric add 
(sp. gr. 1.06 =» 10 per cent. HNOj). As the solution cools, yeUow crystals of 
the nitrate of nitro-papaverine, C2oHjo(NOi)N04.HNO|.HiO, appear. Yellow 
prisms of nitro-papaverine, CioHio(N02)N04.H20, may be obtained from this 
nitrate by means of ammonia. 

4. Anmionia colors the greenish solution of papaverine in 

14 



210 DETECTION OF POISONS 

chlorine water deep red-brown which becomes later almost 
black-brown. 

5. Selenious-Stilphtiric Acid Test — See page 207 for the 
color changes given by pure papaverine dissolved in this 
reagent. • 



PILOCARPINE 

Pilocarpine, C11H16N2OS, occurs with isopilocarpine and probably also with 

pilocarpidine in the leaves of jaborandum (Pilocarpus pennatifolius^). The 

H O free base as usually obtained is semi-liquid, viscous, non- 

CiHi — C — C volatile and alkaline. It dissolves but slightly in water; 

\0 is freely soluble in alcohol, ether and chloroform; and 

HQ Q insoluble in benzene. Solutions of pilocarpine and its salts 

I Hs are dextro-rotatory. This alkaloid is a strong base neu- 

CHi tralizing adds and forming salts that are usually crys- 

y^ ' talline. Caustic alkalies, added to concentrated solutions 

Q }i^ of pilocarpine salts, precipitate the free base which redis- 

\pTT solves in an excess of the precipitant. Solutions of 
1^ ^ sodium hydroxide, or sodium ethylate (CsHf.ONa), cause 

^ a molecular rearrangement of pilocarpine. This reaction 

runs more smoothly, if pilocarpine hydrochloride is heated for half an hour at 
200®. The product of this change is isopilocarpine, CnHieNjOi, isomeric 
and very likely stereo-isomeric with pilocarpine. Both isomeric pilocarpines 
differ in melting points, solubilities and particularly in specific rotation. 
Isopilocarpine is less dextro-rotatory than pilocarpine and crystallizes in 
deliquescent prisms easily soluble in water and alcohol. The salts of the 
two bases also show similar differences: 

Pilocarpine nitrate, CiiHieN202.HNOj; mpt. 178**; [ajo = + 82.90®. 
Isopilocarpine nitrate, CnHieNiOi.HNOj; mpt. 159®; [a]D = 4-35 .68*. 

Jowett' has succeeded in converting isopilocarpine into pilocarpine by means 
of the same reagent used in converting pilocarpine into isopilocarpine. Pure 
isopilocarpine, heated with pure alcoholic potassium hydroxide, gives a mixture 
of unaltered isopilocarpine and pilocarpine. The identity of the latter with 
pure pilocarpine was established by preparing the hydrochloride and nitrate 
(mpt. 178*). This reciprocal conversion of one alkaloid into the other strong^ 
supports the idea of the stereo-isomerism of pOocarpine and isopilocarpine. 
Pinner was the first to show that the two nitrogen atoms of the two isomeric 

^ According to Jowett, jaborine, which has been described as another alkaloid 
peculiar to jaborandum leaves, is a mixture of isopDocarpine, pilocarpidine, a 
little pilocarpine and pigment. 

"Proceedings of the Chemical Society 21, 172 (1905). 



POISONS NOT IN THE THREE MAIN GROUPS 211 

bases belong to a glyoxaline ring.^ In 1905 Jowett proposed for pilocarpine 
and isopilocarpine the following formulae: 

CiH,.CH— CH— CHf— C— N— CH, C,H,.CH— CH— CHr-C— N— CH| 

>CH 



DC 



CH, HC— N DC CH, 



HC— N 



\/ \/ 

O O 

Pilocarpiae Isopilocarpine 



Detection of PUocaipine 

Ether, chloroform or benzene extracts pilocarpine from 
aqueous solutions alkaline with sodium hydroxide or carbonate. 
Evaporation of these solutions leaves a thick, non-crystalline, 
alkaline syrup. The general reagents especially delicate for 
pilocarpine are: iodo-potassium iodide, phospho-molybdic add, 
phospho-tungstic acid and potassium bismuthous iodide. 

I. Place a particle of potassiimi dichromate and 1-2 cc. of 
chloroform in a test-tube. Then add pilocarpine itself, or its 
solution and i cc. of 3 per cent, hydrogen peroxide. Shake for 
several minutes. The mixture yellowish at first gradually 
darkens and in 5 minutes is dark brown. Depending upon the 
amoxmt of pilocarpine, the chloroform is blue-violet, dark or 
indigo-blue. But the upper aqueous solution gradually fades. 
The chloroform mixture is an intense blue from o.oi gram of 
pilocarpine and blue-violet from o.ooi gram and less. The 
color lasts from an hour to a day (H. Helch^). 

Apomorphine (o.oi gram) colors chloroform blue-violet even without hydrogen 
peroxide. Strychnine gives a barely perceptible bluish tint which changes com- 
pletely within a few minutes. There is a color with antipyrine only after 
acidification of the hydrogen peroxide. 

^ Glyoxaline, or imidazole (C1H4N1), is obtained by the action of ammonia 
upon glyoxal in presence of formaldehyde. It is a strong base and crystalline. 

+ + C /" I ^CH + 3H,0. 

HC = iO 3Z 'li^lNiig^^^'^^" TO HC— N 

Glyozol Pormaldehyde Glyoxaline 

' Pharmazeutische Post 35, 289, 498 (1903) and 39, 313 (1906). 



212 DETECnOX OF POISOKS 

2. Mandelin's reagent dissolves jMlocaipiiie with a golden 
yellow color which gradually changes to bright green and finally 
to light brown. 

3. The solution of pilocarpine in formalin-sui^huiic add 
becomes yellow, yellowish brown and blood red, if warmed. 

Thus far fatal poisonings from this alkaloid have not oc- 
curred and nothing is known as to the possibility of its detection 
in the cadaver. 

PTOMAINES 

Ptomames are basic substances contaming nitrogen and may be toxic or 
non-toxic They are produced during putrefaction of cadavers under the in- 
fluence of bacteria. They are to be regarded to some extent as products of 
bacterial metabolism and are nearly always present in cadavers, eapedaDy in 
those parts which are in an advanced state of putrefaction. Jf any ptomaines 
closely resemble alkaloids. Like alkaloids they give precipitates with the gcnenl 
reagents, and certain ptomaines resemble well-defined alkaloids even with apedal 
reagents. Hence ptomaines are of very great importance in forensic chemistry, 
since their presence may easily lead to mistakes and false condusions. These 
putrefactive products also resemble vegetable bases in their behavior with sol- 
vents. Ether extracts some of them from add solution and others from alkaline 
solution, whereas certain ptomaines are removed from alkaline solution only by 
amyl alcohol or chloroform. Most of the ptomaines are strong reducing agents, 
for example, they will immediatdy convert potassium ferricyanide into ferro- 
cyanide. Consequently, they give the Prussian blue test with a dilute mixture 
of solutions of ferric chloride and potassium ferric>'anide. Many of the alkaloids 
like morphine resemble the ptomaines in this respect. 

The resemblance of a ptomaine to a definite vegetable base is frequently con- 
fined to some one reaction, and never extends to all the reactions diaracteristic 
of the particular alkaloid. In a legal-chemical investigation no precaution, 
which guards against mistaking a ptomaine for a vegetable base, should be 
omitted. It is an invariable rule to make every test characteristic of the 
suspected alkaloid, and not to be satisfied with possibly one positive test. A 
determination of the physiological action of the substance should supplement 
the chemical examination. A ptomaine may resemble a vegetable base chemi- 
cally, and yet the two substances may differ very deddedly in ph3rsiological action. 
Thus far, ptomaines have been found which show certain resemblances to coniine, 
nicotine, strychnine, codeine, veratrine, delphinine, atropine, hyoscyamine, 
morphine and narceine. Sdmi has described a putrefactive prcduct uriiich 
resembles morphine. Ether failed to extract it, either from acid or alkaline 
solution, whereas amyl alcohol removed it ^-ith ease from an alkaline or am- 
moniacal solution. It liberated iodine from iodic add, but failed to give the 
tests which are characterbtic of morphine alone, namely, Husemann's, PeUagri's 
and the ferric chloride tests! 



POISONS NOT IN THE THREE MAIN CROUPS 



I 



The object In such c 
doubt. Every poasiblt 
pore state. When this 

established beyond quest! 



must be to get a result about which thete can be no 
ins must be used to isolate the alkaloid in a perfectly 
be accomplished, the nature of the poison can aln'aya 



SAPONINS 



The term saponins, or saponin substances, includes a large number of gluco* 
side-like bodies of widespread occurrence in the vegetable kingdom and having 
in common certain chemical, physical and especially physicological properties. 
Their aqueous solutions when shaken foam readi'y. In this respect they 
resemble the soaps. Many saponin substances have a sharp, harsh taste. In 
powdered form they excite violent sneezing. They arc capable of holding many 
finely divided substances in a state of emulsion. They dialyze incompletely 
»nd salts precipitate them from solution. Eicepling the gluco-alkalcid solanlne, 
which contains nitrogen and is alkaline, the saponins may be classified chemically 
as nitrogen-free glucosidcs. Most saponins are neutral and only a few are 
faintly add. Neutral saponins and alkali salts of acid saponin substances dis- 
solve in water and hot aqueous alcohol but ace msoluble m absolute alcohol and 
ether. Barium hydroxide and lead acetate (neutral and basic) prectpilate 
saponins from concentrated aqueous solution. The former gives baryta saponins. 
Basic lead acetate precipitates all saponins but the neutral salt precipitates 
only add saponins. Ammonium sulphate is capable of salting saponin sub- 
stances from solution as it does proteins. Solutions of saponins in concentrated 
sulphuric acid are yellow, gradually becoming red and sometimes violet and 
blue-green. The detection thus far of saponin substances in more thaif 50 plant 
families having over 300 monocotyledenous and dicotytedenous species shows 
the wide occurrence of these substances in the vegetable kingdom. Saponins 
occur in roots (Senega, Saponaria), tubers (Cyclamen), barks (Quillaja, Guaia- 
cum), fruits (Sapindus, Saponaria), seeds (.'Esculus, Agrostemma, Thea), stents 
(Dulcamara) and leaves (Guaiacua). In fact almost any part of the plant 
organism may contain saponins. The plant families, producing saponin sub- 
stances in greater abundance, are the sapindaceic, caryophyllacen;, colchicacei 
polygalaceie, Hdencie and solanacei. Quite considerable quantities of saponins 
may occur in the particular part of the plant. 

Saponin solutions, heated with dilute hydrochloric or sulphuric add, are 
hydrolyzed into sugars and a non'toxic substance msoluble in water called 
sapogenin. The sapogenins have not been extensively investigated but they 
ate not entirely identical. 

The following saponins have been mote dosely stucUed: 
I Digitonin; in the seeds of Digitalis purpurea. 
I Saponin: in the root of Saponaria otiictnalis (4-5 pet cent.). 
I Githagin: in the seeds of the corn cockle, Agrostemma githago (6,5 per 
cent.). 

Senegin: i: 

Struthiin: 
cent). 

Quillaja-Sapc 



Senega root, the n 
a levantine soap n 



t of Polygala senega. 

t, the toot of Gypsophila struthium (14 per 



n the bark of Quillaja saponaria (S.S per c< 



DETECTION OF POISONS 



Sapindus-Sapotoxin: i 
Sarsaparilla-Saponin : 
mulan. 



the fruit of Safdndus saponBria. 



e saisaparilla 



, the toot of % 



s kinds of 



Pbysiolosical Action of Saponins. — Almost without exception saponin sub- 
stances are highly toxic, if introduced dirccUy into the blood. Most saponins 
are absorbed with difficulty. Consequently healthy individuals may take dilute 
saponin solutions by the mouth in considerable quantities without ill effects. 
Toxic saponins act in common as protoplasmic initants. In larger doses saponin 
substances kill protoplasm. They manifest in various ways their power of acting 
as protoplasmic poisons. Saponins act upon blood- corpuscles for the same 
reason. S.. Koberl and his collaborators have shown defibrinated blood, diluted 
loo times with physiological salt solution (see below), to be the best and most 
convenient reagent for saponin substances. Saponins cause hxmolysis and the 
blood solution becomes laky. Agglutination and fotnation of methzmoglobin 
do not occur. The freer the blood is of serum, the more pronounced the he- 
molytic action of saponin substances upon blood- corpuscles. Recent investi- 
gations have shown that saponins act more vigorously upon blood- corpuscles 
isolated from serum, because blood serum contains cholesterin which has a pro- 
tective influence and retards haemolysis. Most likely the hemolytic action of 
saponins is due to removal of cell membrane lecithin, the chief constituent of the 
cell wall, from red blood- corpuscles, for lecithin -saponins are formed. Saponins 
also combine with cholesterin, us well as with lecitliin, fotming cholcstcrin- 
saponins. The affinities of a saponin having been satisfied by cholesterm, it no 
longer acts upon the lecithin of the membrane of blood- corpuscles. Thus 
cholesterin prevents hiemolyais, which a saponin may produce, and so acts as 
an antidote to saponin substances. Ransam' bos made the important discovery 
that addition of cholesterm checks the solvent action of a saponin upon blood- 
corpuscles. At flrst it was not known whether this antidotal action was due to a 
chemical reaction, or to adsorption, that is to say, lo a physical process. R. 
Eobert' as well as Madsen and Nogucbi' were able to dissolve cholesterin, 
which is insoluble in water, m an aqueous saporun solution. They assumed that 
this physiologically inactive solution contained a labile saponin-cholesterin 
compound no longer having bicmolytic power. Recently A. Windhaus* has 
definitely proved that saponin-cholestcrides exist. Digitonin-cholesteride, 
CuHmOii.CjvHisO, crj-stallizes in fine needles, when a hot alcoholic solution of 
digitonin (r molecule) is poiucd into a similar solution of cholesterin (i mole- 
cule). This cholcsteride is formed without elimination of water. Hence in this 
reaction between digitonin and cholesterin we ate dealing most probably with the 
formation of a molecular compound. 

Saponin solutions also dissolve white blood- corpuscles but only at higher 
concentrations. A physiological action characteristic of many saponins ii 
exhibited in the stupefaction and killing of fish, even in water containing only 



:3oo,ooo of saponin substance (R. Kobert). 

' Deutsche medizinische Wochenschrift igor, 194. 

* R. Kobert, Die Sapanine, Stuttgart, 1904. 

* Chemisches Zentralblalt, 1905 I, 1:65. 

' Berichte der Deutschcn chcmischcn Gescllschaft 42, 



^^1 



POISONS NOT IN THE THKEE MAIN GROUPS 215 

Detection of Saponins 

The matter of solubility is especially important in isolating 
iaponin substances from mixtures. All saponins are soluble in 
water and some in alcohol, but they are practically insoluble 
in ether, benzene, chloroform and petroleum ether. Employ 
neutral or basic lead acetate (see above) in isolating saponins. 
Decompose the washed precipitate with hydrogen sulphide, 
filter and evaporate the filtrate upon the water-bath. Pre- 
cipitate the saponin with absolute alcohol and ether from the 
concentrated solution. Solutions of most saponins in con- 
centrated sulphuric acid are red or yeUowish red, gradually 
becoming violet. Saponin substances give various colors with 
Froehde's reagent and vanadic-sulphuric acid: brown, red- 
brown, blue, green and violet (see solanin). A saponin solution, 
heated with dilute hydrochloric acid, undergoes hydrolysis and 
then, owing to formation of sugar, reduces FehUng's solution 
with heat. 

»Dete 
Treat the beverage to be tested for saponin with excess of 
basic magnesium carbonate, evaporate to about too cc. and mix 
with 2 volumes of 96 per cent, alcohol. Filter after 30 minutes 
and evaporate the alcohol from the filtrate. Filter the residue 
hot and extract the cold filtrate with sufficient liquid carbolic 

•acid' to leave about 5 cc. undissolved. Add ammonium sulphate 
to hasten the separation of the carboUc acid layer. Then shake 
the latter with water and a mixture of 2 volumes of ether and i 
volume of petroleum ether. Evaporate the aqueous solution 
to dr>'ness upon the water-bath. Wash the residue with cold 
absolute alcohol, in case of wine, and with acetone, in case of 
The residue fails to give the saponin reaction well, 
ihat is to say, a red color with concentrated sulphuric acid, un- 
!ss treated as described. E. Schaer dissolves the residue in con- 

* E. BruitneT, Zeitschrift fQr Unteisudiung der Nahrungs- und Genussmittel 
I "97 (1902). 
■ Addum carbolicum liquefactum of the German PharmBCopceia. 



detection in Foaming Beverages (Beer, Wtne, Effervescing Lemonade) ■ 



216 DETECTION OF POISONS 

centrated aqueous chloral hydrate solution and adds the latter 
to concentrated sulphuric acid as an upper layer. A saponin 
produces a yellow, then purple-red and finally mallow-blue 
zone. 

Detection of Githagin (Com Cockle Saponin) in Flour 

Heat 500 grams of flour with i liter of alcohol (sp. gr. 0.8496 
= 85 per cent, by volume). Filter hot, distil most of the 
.alcohol, add absolute alcohol as well as ether to the residue and 
let stand 12-24 hours. Collect the precipitate upon a filter and 
dry for a short time at 100° to coagulate possible protein. 
Dissolve in a little cold water, filter and precipitate githagin 
from the filtrate with absolute alcohol, best with addition of 
ether. Githagin thus obtained is a yellowish white powder 
having a sharp, harsh taste. 

To prove the presence of a saponin substance, agitate its 
aqueous solution which should foam. Then heat the solution 
with dilute hydrochloric acid and test its reducing power with 
Fehling's solution. Finally, if possible, perform the physio- 
logical test with blood. Dilute defibrinated ox blood with 100 
volumes of 0.9 per cent, sodium chloride solution and add 
the solution of supposed githagin in 0.9 per cent, sodium chloride 
solution. The blood solution at once becomes laky, if githagin 
is present. According to J. Brandl,^ Agrostemma-Sapotoxin 
(githagin) produces haemolysis in very great dilution (i : 50,000). 
But after previous treatment with cholesterin, even o.oi gram 
shows no haemolytic action whatever. 

Physiological Salt Solution and Haemolysis 

To prevent red blood-corpuscles from changing volume in 
experiments requiring dilution of blood, an isotonic salt 
solution must be used. What is an isotonic solution? If n 
gram-molecules of a body A are dissolved in a definite volume of 
solvent and n gram-molecules of a body B are dissolved in an 

^ Archiv £Ur experimentelle Pathologic und Pharmakologie, 54, 245. 



POISONS NOT IN THE THREE MAIN GROUPS 



217 



I 

I 
I 



equal volume of the same solvent, certain properties of the 
original solvent are changed equally in both cases. The freezing 
point of the solutions is lowered and the boiling point raised 
equally. The two solutions have the same vapor tension and 
the same osmotic pressure. In other words they are isotonic. 
Blood-corpuscles retain their volume unchanged, if brought 
into a salt solution having the same osmotic pressure as the 
blood serum. Such a salt solution is isotonic with blood serum. 
In the case of human and mammalian blood an isotonic solution 
of sodium chloride has a concentration of 9 per thousand = 
physiological salt solution. Such a solution formerly contained 
0.6 per cent, of sodium chloride. Blood-corpuscles give up 
water to solutions of higher concentration than 0.9 per cent. 
NaCI (hyperisotonic solutions) until osmotic equilibrium is 
established. They shrivel and hence have a smaller volume.. 
On the other hand, blood-corpuscles in salt solutions of lower 
concentration {hypisotonic solutions) take up water and become 
distended. In diluting blood with water, this swelling may 
go far enough to cause hemoglobin to separate from the stroma 
and pass into the aqueous solution. This process is called 
hjemolysis. Alternate freezing and thawing of blood may 
produce hsemolysis. Various chemical substances, which act as 
protoplasmic poisons, cause the same result. Such substances 
are ether, alcohol, chloroform, alkalies, gallic acids, solanine, 
etc. The saponins described above are also powerful hsemolytic 
agents. Finally, those globulicidal substances, or haemolysins, 
normally occurring in blood sera, as well as those produced in 
immunization, belong in this class. 

SOLAHINE 

Solanine, CtiHgjNOn, al the some time an alkaloid and a glucoside (glueo- 
alkaloid] occurs in the potato plant ISolanum tuberosum) and in other Solanacex 
u Solanum nigrum, Solonum dulcamara and Soianum lycopcisiciun ([omato). 
It has been found also in Scopoliaccie, aji in ScopoJia orientolis and ScopoUa 
Atropoides. Solanioe is not uniformly distributed in all pacts of the potato plant 
but ia most abundant in the berry-like fruit and m the chlorophyll- free sprouts 
appearing m the spring upon potatoes that lie in a cellar. Schmiedeberg and 
Ueyer found 0.034 gram of solanine per Idlogram of peeled potatoes in January and 



218 DETECTION OF POISONS 

F^ruary bat 0.044 gram in tiiq>eeled potatoes. Potato pedmgs gave 0.71 gnun 
of solanine per kilogram and potato sprouts i cm. long even sx> grams. The 
^>pearance of solanine accordmg to R. Weik is due to the life prooesses of Bac- 
terium scJaniferum (?). 

Sc^anine crystallizes in white needles having a bitter taste and melting at 
344''. Even boiling water dissolves only a little of this alkaloid (about i: 8000). 
It is sdiuble in 500 parts of cM and 125 parts of b<Mling alcohol; and in about 
4000 parts of ether. These solutions are faintly alkaUne. Hot saturated sohitiooB 
of solanine in alcoh<d and amyl alcoh<^ gelatinize upoa cooling. Ether, cfahxo- 
form and benzene do not extract solanine either from add or <^iv^lw*^ iwJutwti 
But hot amyl alooh<d extracts solanine from aad solution and from solutioni al- 
kaline with sodium hydroxide or ammonia. Solanine is a weak base, readily db- 
8<^ving in adds, as acetic add, and forming crystalline salts. Dilute hydrochloric 
or sulphuric add hydrolyzes solanine to solanidine, C40HtiKOs, galactose and 
rhamnose. Hydrolysis is very slow m the cold but rapid upon heating. The 
hydrochloride or sulphate of solanidine separates as a difficultly soluble, crys- 
talline powder. A good yield of s<^anidine is obtained, according to Wittmann, 
by heating solanine under a return-condenser with 10 times the quantity of 2 
per cent, sulphuric add, until the liquid is yeUowish and the filtrate upon further 
boiling no longer deposits solanidine sulphate. Solanidine, precipitated from its 
sulphate with ammonia and recrystailized from ether, forms colorless, silky 
needles, melting at 207** and dissolving with difficulty in water but readily in ether 
or hot aloohoL Solanidine is a stronger base than solanine and the salts it forms 
with adds are usually crystalline and difficulty soluble in water. Solanine and 
solanidine are highly toxic substances having an action similar to that of the 
saponin substances (see above). 

Toxic Action. — Solanine taken internally is usually very imperfectly ab- 
sorbed. As a glucoside its action is local and as a saponin-like substance strongly 
hemolytic, rendering the blood laky. A solanine solution even in a dilution 
of 1 : 8300 causes complete hemolysis. Internal administraticm of solanine usually 
produces emesis and larger doses cause gastro-enteriUs (gastro-intestinal catarrh). 
The latter also f oUows intravenous and subcutaneous injecticm of doses not rapidly 
fatal At the same tmie a hsmoglobinuna may appear. (R. Kobert, In- 
toxikationen). 

Detection of Soknine and Sdanidine 

Since very dilute mineral adds hydrolyze solanine, these adds 
cannot be used to detect this alkaloid. E. Schmidt^ suggests 
the following procedure. Extract the material with cold water 
containing tartaric add. Neutralize the filtered extract with 
calcined magnesia and evaporate to dryness upon the water- 
bath. Extract the residue with alcohol and filter hot. If the 

1 Pharmazeutische Chemie, Organischer TdL 



POISONS NOT IN THE THREE MAIN GKOUPS 



219 



quantity of solanine is not too small, the alcoholic extract gel- 
atinizes upon cooling. Otherwise, evaporate the alcoholic 
solution and examine the residue for solanine. L. Robert 
extracts solanine with isobutyl alcohol from alkaline solution. 
Phospho-molybdic acid is the only general reagent giving a pre- 
cipitate with a solanine solution and that is yellow. But sol- 
anidine, that is to say, a solanine solution that has been boiled 
with excess of hydrochloric acid, being a stronger base, gives 
precipitates with most of the other general reagents. 

Special Tests for Solanine and Solanidine 



1. A solution of solanine in selenic-sulphuric acid' is rasp- 
berry red. Gentle heat favors the appearance of this color. 
Solanidine gives the same result. 

2. Solutions of solanine and solanidine in vanadic-sulphuric 
acid' are orange-yellow, soon becoming red and finally blue- 
violet. Solanine may be dissolved first in sulphuric acid and a 
drop of vanadic-sulphuric acid added to this solution. 

3. Solutions of solanine and solanidine in ethyl-sulphuric acid* 
are red. An alcoholic solution of solanine, carefully added to 
concentrated sulphuric acid as an upper layer, produces a red 
zone where the two liquids meet (E. Schmidt). 

4. A solution of solanine in concentrated sulphuric add is 
orange but becomes brownish red on longer standing or gentle 
wanning. Red streaks appear, if bromine water is added drop 
by drop to a solution of solanine in concentrated sulphuric 
acid. 

5. A solution of solanine in Froehde's reagent is first yellow- 
ish red then evanescent cherry red and finally red-brown. 

The methods for estimating solanine quantitatively in pota- 
toes are described in Chapter VI (see page 284). 

' A mixture of 1.3 grams of sodium aelenate (NajSeOt.io HjO), 8 tc. of water 
and 6 cc. of concentraLcd sulphuric acid. 

' Dissolve o.i gram a{ ammomum vanadate (HiN.VOi) in 100 grams of concen- 
trated sulphuric acid. 

' Add 6 cc. of concentrated sulphuric add to 9 cc. of absolute alcohol. 



I 



I 



220 DETECTION OF POISONS 

THEBAINE 

Thebaine, CigHnNGi - CiifIii(OCHs)iNO, constitutes about 0.15 per cent. 

of opium. This alkaloid crystallizes from dilute alcoh(d in leaflets having a 

XT TT silvery glitter and from absolute alcohol in prisms 

C C N.CHi n^clting at 193**. It is nearly insoluble in water, 

^\/\/\ rather easily soluble in hot alcohol, ether, benzene 

HC C CH CHj and chloroform. It differs from morphine in being 

CH O C C C CH ^^^^y u^soluble in caustic alkalies. Its soluticms are 

' \/\^\/ tasteless and laevo-rotatory. 

C C CH Constitation;— Thebaine is a strong tertiary base, 

I I i forming as a rule well crystallized salts with adds. 

V^ ^^^ excess of add, espedally mineral add, usually 

Q decomposes these salts with ease. Being a tertiaiy 

base, it easily combines with methyl iodide, forming 



„ „ ^ , 9^^*, thebame lodomethylate, CisHhNGs.CHiI, crystalUz- 

R. Pschorr s formula ... r« * .1 .1 . 

mg m pnsms. Two of the three oxygen atoms m 
thebaine are methoxyl-groups ( — OClls) and the third probably forms an 
ether-like combination, a so-called bridge-oxygen. The thebaine molecule 
appears not to contain hydroxyl. 

Heated with acetic anhydride, thebaine gives the acetyl derivative of the 
phenol thebaol, CieHuOi, and a nitrogenous product, methyl-oxy-ethylamine, 
CH9.NH.CH2.CH2.OH. R. Pschorr has S3mthesized thebaol, or the methyl 
ether of thebaol, and shown by this synthesis that thebaol is 3,6-dimethoxy- 
4-oxy-phenanthrene (see bdow). Pschorr assigns to thebaine the structural 
formula given above which is analogous to that of apomorphine and of morphine 
(see pages 1 22 and 126). Thebaol has the foUowing structural formula: 

(i)H H 
C C 

^\/\ 
HC C CH 



(3)CH,0.O C C 



C C CH 



(4)H0 HC CH 
C 
0CH,(6) 

Detection of Thebaine 

Ether and chloroform extract thebaine from an alkaline 
aqueous solution and consequently this alkaloid appears in 
ether extract B, if the Stas-Otto procedure is followed. The 
general reagents, phospho-tungstic acid, iodo-potassium iodide, 
potassium mercuric iodide and potassium bismuthous iodide 



POISONS NOT IN THE THREE MAIN GROUPS 221 

precipitate thebaine even from very dilute solutions. Thebaine 
gives the following color reactions: 

1. Conceiitrated Sulphuric Acid gives a deep red color with 
thebaine and the solution gradually becomes yellowish red. 
Froehde's reagent gives the same result. 

2. Concentrated Nitric Add dissolves thebaine with a yellow 
color. With Erdmann's reagent the color varies from dark red 
to orange. 

3* Chlorine Water dissolves thebaine and ammonia turns the 
solution an intense red-brown. 

Tozalbumins 

Toxalbiunins are toxic, protein-like substances either already formed in the 
plant or animal organism, or produced in the metabolism of pathogenic micro- 
organiskns. These substances as yet have not been isolated pure as individual 
chemical compounds. The chemical and physiological properties of such 
vegetable toxalbumins as abrin, ridn, robin and crotin are given as a matter of 
fact by substances obtained from some particular part of the plant by a definite 
method. The vegetable toxalbumins mentioned possess the common property 
of clumping, agglutinating and precipitating red blood corpuscles. Therefore 
R. Kobert classifies them as ''vegetable agglutinines." A trace of one of these 
agglutinines, added to defibrinated blood in a test-tube, causes clumping into a 
mass resembling sealing-wax. Abrin, ricin and crotin also cause coagulation of 
milk. 

Abrin 

This toxalbumin occurs in jequirity seeds from Abrus precatorius. Remove 
the seed envelopes and extract the finely divided seeds with 4 per cent, sodium 
chloride solution. Concentrate the filtered liquid in vacuo and addify with 
acetic acid. Precipitate abrin from this solution by addition of sodium chloride 
and finally purify by dialysis. Abrin is an amorphous, highly toxic powder not 
entirely free from ash. Though abrin and ricin are alike in some respects, they 
are not identical. 

Ridn 

This intensely toxic toxalbumin constitutes 2.8-3 P^ ^^^^ ^^ ^^^ castor bean. 
Remove the seed envelopes and subject the seeds to powerful pressure to remove 
as much oil as possible. Then extract with 10 per cent sodium chloride solution. 
Saturate the filtered extract at the same time with magnesium and sodium sul- 
phate and keep for some time in the cold at room temperature. Place the pre- 
cipitate, which contains ridn, in a parchment paper dialyzing tube and dialyze 
for several days. Finally dry the ricin left in vacuo over sulphuric add. Ridn is 



222 DETECTION OF POISONS 

an amorphous, highly toxic powder containing ash and easily soluble In lo per 
cent, sodium chloride solution. ThiS toxalbumin, dissolved in sodium chloride 
solution, gives the protein reactions. Ridn possesses in high degree the power of 
agglutinating blood corpuscles. Use defibrinated blood for this test-tube experi- 
ment, not diluted blood or blood mixed with physidogical salt solution. Riciny 
according to Elfstrand, agglutinates the red blood corpuscles of the guinea-pig 
even in a dilution of i : 600,000. Ridn agglutinates the blood of all mammals but 
not to the same degree. Removing serum from the blood and substituting 
physiological salt solution strengthens rather than weakens the agglutinating 
action of ridn. The inference is that serum must have a certain anti-agglu- 
tlnating action. Separation of red blood corpusdes into stroma and hemo- 
globin^ shows that ricin has not changed hemoglobin in the least. But the 
stromata have been altered just as the blood corpusdes have been. 

To detect ridn in castor bean press -cake, or in feeds containing castor beans, 
extract the findy divided material with physiological salt solution at room tem- 
perature, filter and make the agglutination test in a test-tube with imdiluted, de- 
fibrinated blood and with blood diluted with physiological salt solution. 



Crotin 

Crotin is a substance obtained from the seeds of Croton Tiglium. Remove the 
seed envdopes, express the oil and treat as described for abrin and ridn. Chem- 
ically crotin is very similar to ricin. Abrin and ridn agglutinate the blood cor- 
pusdes of all warm-blooded animals thus far tested but crotin does not behave 
the same with all kinds of blood. (See R. Kobert, Intoxikationen.) 

Coagulation of Blood and Defibrinated Blood 

Blood is a transparent fluid, the blood plasma, suspended in which is a very 
large number of solid partides, the red and white blood corpusdes. Outside 
the organism blood coagulates even in a few minutes after being drawn. In the 
dotting of blood a very difficultly soluble protein, called fibrin, separates. If the 
blood is still, the dot is a solid mass which gradually contracts and exudes a dear 
liquid, usually yellow, the blood serum. The coagulum, thus formed and envdop- 
ing the blood corpusdes, is called the crassamentum (Placenta sanguinis). But 
if the blood is whipped during coagulation, fibrin separates in threads. The 
fluid separated from the latter is defibrinated blood which consists of blood cor- 
puscles and blood serum. To obtain defibrinated blood, whip the fresh blood 
removed from a vein with twigs and fibrin will separate on these. Or run the 
fresh blood into an Erlenmeyer flask, containing iron filings, and shake vigorously 
for several minutes. Fibrin is predpitated on the filings. 

There are several ways to retard coagulation of blood, among which the follow- 
ing may be mentioned: 

I. Cool blood suddenly to low temperature. 

^ The two prindpal components of blood corpuscles are the stroma, which con- 
stitutes the true protoplasm, and the intraglobular contents, the chief constituent 
of which is haemoglobin. 



POISONS NOT IN THE THREE MAIN GROUPS 223 

3. Draw blood direct from the vein into a netural salt solution, for example, 
magnesium sulphate solution (i volume of salt solution and 3 volumes of blood) 
and stir. This mixture of blood and salt will not coagulate for a day. 

3. Add blood to sufficient dilute potassium oxalate solution to give a mixture 
containing 0.1 per cent of oxalate. The soluble calcium salts of the blood are 
precipitated by the oxalate and the blood loses its power of coagulating. 

4. To prepare a non-coagulating blood plasma, poiu* blood into sodium fluoride 
solution until it contains 0.3 per cent of NaF. 



CHAPTER V 
SPECIAL QUALITATIVE AND QUANTTTATIVB METHODS 

Quantitative Estimation of Phosphorus in Phosphorated Oils 

I. W. Straub's Method. — Straub has found that his test^ with 
dilute copper sulphate solution, recommended for the qualita- 
tive detection of phosphorus, may also be used to determine 
phosphorus in a phosphorated oil. If such an oil is shaken with 
3 per cent, copper sulphate solution, there is first a brownish 
black emulsion in which each individual oil drop is coated with a 
film of copper phosphide, PCua (?). After 4-5 hours shaking, 
this brownish black colol disappears and the mixture separates 
into two layers. All the phosphorus in the oil is now in the 
aqueous solution as phosphoric acid. This method has the 
further advantage that the decolorization of the emulsion serves 
as an indicator of the completion of the oxidation. 

Procedure. — Put 25 cc. of 3 per cent, copper sulphate solution 
(taken as CUSO4.5H2O) in a separatory funnel. Add 5 cc. of 
the phosphorated oil* and agitate the mixture vigorously for a 
long time. If a shaking machine is available, place the mixture 
in a thick-walled glass bottle with a tight glass stopper and shake 
3-5 hours, or until the original brown emulsion has disappeared 
and become clear and bright blue. Separate the aqueous solu- 
tion in a separatory funnel and precipitate phosphoric acid at 
once by the molybdate method and finally weigh as magnesium 
pyrophosphate, Mg2P207. 

^ Zeitschrift fUr anorganische Chemie 35, 460 (1903). 

' To prepare a phosphorated oil suitable for such determinations, dissolve about 
0.1 gram of yellow phosphorus in the smallest possible quantity of warm carbon 
disulphide and dilute this solution to 100 cc. with olive oil. Although carbon 
disulphide does not affect the determination of phosphorus, it may be removed 
by warming the phosphorated oil on the water- bath. 

[224 



SPECIAL QUALITATIVE AND QUANTITATIVrE METHODS 225 



—The accuracy of this method is shonn by the tesulta of Suaub's 
deteccainatiooB. Instead of 0,005 S^^^' of phosphorus, dissolved in 5 cc. of oil, 
he found 0.0047 aQ() 0.00468 gram. Even very considerable dilutions of the 
phosphorated oil do not affect the accuracy of the determination, la the case of 
the more concentrated phosphorated oils, shaking with copper sulphate solution 
must be kept up much longer. 

2, A. Frankel's^ and C. Stich's^ Method. — Dissolve the oil 
in acetone and precipitate phosphorus with hot alcoholic silver 
nitrate solution. Oxidize the phosphorus in the precipitate to 
phosphoric acid and finally determine the latter in the usual 
way. 

Procedure. — Dissolve 20-50 cc. of the phosphorated oil, as 
phosphorated cod liver oil, in 100 cc. of acetone or ether and 
completely precipitate with hot alcoholic silver nitrate solu- 
tion.' First wash the precipitate of silver phosphide with 
ether-acetone mixture and then with alcohol. Treat next with 
hot 25 per cent, nitric acid, containing a little fuming acid. 
Expel excess of nitric add from the filtrate on the water-bath 

id precipitate silver with hydrochloric acid. Finally filter 
■om silver chloride and determine phosphoric acid in the filtrate. 



Ex 

Bro 



Remarks. — Since sodium hypophosphite and phospliite are soluble in acetone 
and also precipitated by acetone-silvec nitrate, it is advisable first to extract a 
test portion of the phosphorated oil with water and then test the aqueous extract 
for these first oxidation products of phosphorus, hypophosphorous and phosphor- 
ous adds. If they are present, all the phosphorated oil should first be eibacted 
with water in the same manner. 

Phosphorus in phosphorated oils, especially phosphorated cod liver oil stowty 
disappears. C. Stich found that a phosphorated cod liver oil, containing 0.05 
per cent, of phosphorus, mth the usual daily removal of 5 grams, lost in j weeks 
only 3-5 miUigrams of phosphorus. Such a decrease in the amount of phosphorus 
in phosphorated oils is only of slight significance. Dilute oily solutions of phos- 
phorus (1:1000), when kept in tightly stoppered boltlesand protected from light, 
are constant as regards their phosphorus content for a long time, even 5-6 months. 
Moreover, phosphorous much diluted as vapor or in solution, is oxidized with 
corresponding dilSculty. The same is also true of phosphorus in the animal or- 
ganbm. Therefore it is possible sometimes to detect free phosphorus in the 
excretory organs, as the liver, even several weeks after phosphorus posioning. 

The distillation method is inapplicable in the quantitative estimation of phos- 

> PharmaJieutische Post 34, 117. 

* Phannaieutische Zcitung 37, 500 (1901). 

ate dissolves in about to parts of alcohol. 



226 DETECTION OF POISONS 

phorus in oils, as cod liver oil, since only about 40 per cent of the phoqkhonis 
present is found in the receiver, even when the strongest 'oxidizing agent and the 
best absorbent for phosphorus are used. To place the phosphorus-content of the 
cod liver oil residue at the amount of the distilled phosphorus is no^ admissible, 
because cod liver oil as such contains about 0.02 per cent, of combined phosphorus. 

Special Methods for the Detection of Arsenic 
Isolation of Arsenic as Arsenic Trichloride^ . 

This depends upon the volatility of arsenic as chloride, AsCU, 
in concentrated hydrochloric add solution and in presence of 
ferrous chloride. The latter serves (a) to reduce any arsenic 
acid possibly present in the material to arsenious add which with 
concentrated hydrochloric add then forms arsenic trichloride 

(a) HiAsOi + 2HCI + aFeClj = H,AsOi + H,0 + aFeCli, 
03) H,AsOi + 3HCI = AsCli + 3H,0. 

Procedure. — Comminute the material and mix with very 
concentrated hydrochloric add (about 40 per cent.) until rather 
thin. Then add 5 grams of ao per cent, arsenic-free ferrous 
chloride solution or saturated ferrous sulphate solution and 
put the mixture into a capadous retort, the neck of which is 
directed obliquely upward and connected with a Liebig cooler 
by an obtuse angle tube, and carefully distil. Distil about a 
third to a half of the original mixture. Dilute the distillate 
with water and test for arsenic in the Marsh apparatus, using 
hydrochloric acid for the evolution of hydrogen. 

If a tubulated retort is used for the distillation, hydrochloric add gas can be 
passed in during distillation so that the liquid being distilled is kept saturated with 
this acid. 

Electrolytic Detection of Arsenic 

To detect arsenic electrolytically, put the liquid, as the sul- 
phuric add solution obtained according to thegeneral procedure 
which contains arsenic as arsenic add^(see page 150), or urine or 
stomach contents, in a suflBidently wide U-tube with platinum 
electrodes (Fig. 18). Pass the current through the liquids 

1 H. Beckurts, Archiv der Pharmazie 322, 653 (1884). 



SPECIAL QUALITATIVE AND QUANTITATI\*E METHODS 227 

acidified with sulphuric acid, and arsine, AsHa, together with 
hydrogen will appear at the cathode, if the liquid contains ar- 
senic. First test the hydrogen for arsenic by the Gutzeit arsenic 
test (see page 1 56) , If a yellow spot appears on the paper moist- 
ened with saturated silver nitrate solution, arsenic is present. 
That this is actually arsenic may be shown by connecting the U- 

Itube as shown in the sketch with a chloride of calcium tube and 
ft Marsh reduction tube; an arsenic mirror then appears in 




Fig, 18. — Apparatus for the ElecLrolytic Detection of Arsenic. 

the latter when heated to redness. Use a current having an 
electromotive force of 7-8 volts. The electrolytic method is 
especially adapted for the detection of arsenic in inorganic 
compounds present in secretions, as the urine, but not for arsenic 
in organic combination as cacody! compounds and arrhenal. An 

Ifiiception among these organic compounds of arsenic is atoxyl, 
pr the anilid of meta-arsenic acid, AsOi.NH,CgH(. The arsenic 
being rather loosely bound is broken up by the electric current 
irith formation of arsine. 



Destruction of Organic Matter and Detection of Arsenic 

(According to A, Gautier' and G. Lockemann') 



This method is of scientific interest rather than of practical significance in 

> Bulletin dc la Sod£t£ diimique de Paris, ig, 639 (1Q03). 

■ Z^tsduift filr angewandte Chemie 18, 416, 491 (ipoj); also ig, 13^3 (igoG). 



228 DETECTION OF POISONS 

forensic chemistry. The purpose is to iQcrease the delicacy of the Marsh- 
Berzelius test for arsenic, and to exclude as far as possible sources of error con- 
nected with the destruction of organic matter, the precipitation of arsenic with 
hydrogen sulphide and the evolution and drsring of hydrogen gas. Organic 
matter is destroyed without the use of hydrochloric add, and arsenic is detected 
without precipitation as arsenic sulphide. Lockemann recommends the following 
procedure and uses finely divided meat as a test experiment: 

Place 20 grams of finely chopped meat in a porcelain dish and add a few cc 
of a mixture to 10 parts of fuming nitric add and z part of concentrated sulphuric 
add« Warm upon the water-bath. The action of the add mixture is so vigorous 
that, even after the addition of about 5 cc, the entire mass, which puffs up con- 
siderably at first, changes to a yellowish, homogeneous, thick, oily liquid. If too 
much add b added at once during warming upon the water-bath, the action may 
be violent enough to cause sudden charring of the whole mass with copious evolu- 
tion of smoke. Such an occurrence may result in loss of arsenic Consequently, it 
is advisable to add the add mixture, amounting in all to' about 20 cc, to the meat 
in 1-2 cc. portions, not adding a fresh portion of add imtil brown fumes cease 
coming ofiF. The mass is dark yellow and finally becomes brown after long heat- 
ing upon the water-bath. Stir with a concentrated aqueous solution of 20 grams 
of a mixture of potassium and sodium nitrate (i -f i) and evaporate upon 
the water-bath. There remains a yellow, crystalline residue which stLU contains 
organic matter. Gradually introduce this mixture in small portions into a 
platinum crucible containing 10 grams of fused potassium and sodium nitrate 
(i -f i). Having added all the mixture, heat the crudble for a short time over 
a free flame. Dissolve the cold melt in water, add sulphuric add and heat upon 
the water-bath until nitrous fumes have been expelled. Test a cold solution of 
the residue for arsenic in the Marsh apparatus. 

Lockemann formerly predpitated arsenic with aluminium hydroxide, Al(OH)s. 
Add 10 cc. of a 1 2 per cent, solution of crystallized aluminium sulphate, AltCSOOr 
18H3O to the solution of the melt free from carbon dioxide and nitrous add. 
Render the solution alkaline with ammonia and heat about 30 minutes upoii the 
water-bath. Collect the predpitate upon a paper, wash with water containing 
ammonia and dissolve in about 30 cc. of 10 per cent, sulphuric add. Heat the 
solution in a porcelain dish upon the water-bath untU it no longer gives a test for 
nitric add with diphenylamine-sulphuric acid.^ Then examine this solution for 
arsenic in the modified Marsh apparatus^ devised by Lockemann (Fig. 19). 

Lockemann's latest results have shown that ferric hydroxide is much more 
effective than aluminium hydroxide as a predpitant of small quantities of arsenic 
Render the water solution of the melt (see above) slightly add with sulphuric 
acid, add a few cc. of iron alum solution, then in the cold, best after cooling with 
ice, add just enough ammonia to predpitate all the iron. Filter after 30 minutes, 
wash the predpitate with cold water to remove nitrates completdy, then dissolve 

* Dissolve I gram of diphenylamine in 100 grams of concentrated sulphuric add. 
A drop of the liquid with a drop of this diphenylamine solution in a porcelain dish 
should not give a blue color. 

* O. Pressler, 30 Bruederstrasse, Leipzig, Germany, supplies this apparatus and 
also the ignition tubes. 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 



229 



n dilute sulphuric acid and test the solution for arsenic in the Marsh apparatus. 
"ton salts do not interfere with the delicacy of the Marsh test for arsenic. 
Zinc in sticks* and sulphuric add are used in the preparation of^bydrogen. 
"topper is the beat activator of zinc in the Marsh apparatus. Break the,iinc 
sticks into pieces weighing about j.i-i.B grams, place for a nuauteino.spMcenL 
copper sulphate solulioa, wash with water, dry with filler paper and preserve care- 
fully in a dosed bottle. This procedure does not interfere with the formation of 
the mirror, whereas addition of copper sulphate to the reduction flask c 
^^ention of arsenic. Copper sulphate used for this purpose should be carefully 




purified by several recrystallizations. The basic properties of fused and granu- 
lated calcium chloride, which are not entirely removed even by hydrogen chloride 
and carbon dioxide, make this an unsuitable drying agent for hydrogen. Locke- 
mann found that potassium carbonate, phosphorus pentoxidc and concentrated 
sulphuric acid cause a noticeable decomposition of arsine. and the same is L 
glass wool and cotton. Crystallized calcium chloride in pieces about i cc. in 
volume is thebest drying Bgent,because it isentirely indifferent to arsine. Locke- 
mann's special drying tube (see sketch) is adapted for the use of this substance. 
Bohemian glass, having a wall thickness of i mm. and an internal diameter o: 
4 mm., is used for ignition tubes. These are drawn out in two places to a length of 
4 cm. The outer diameter of the constriction is t.5 mm. and tlic inner about o. 
mm. The reduction Qask contains 4-6 pieces of coppered zinc and about 15 ci 
of 1 5 per cent, sulphuric acid are added from the dropping funnel. After hydrt 
gen haa been passing through the apparatus for 30 minutes, heat is applied in front 

' Lockemann has found Knhlbaum's stick zinc always arsenic-free. The 
taxae may be said of Bertha spelter from the New Jersey Zinc Company. 




230 DETECTION OF POISONS 

of the first constriction of the ignition tube. If the materials are arsenk-free 
after 1.5-2 hours heating, place the flame in front of the second oonstrictkni 
of the ignition tube. The solution of the iron hydroxide prec^iitate, prepared as 
described above, is added to the reduction flask from the dropping funnd which is 
washed with a little water or dilute sulphuric acid. In testing for very small 
iquantities of arsenic, it is advisable to cool the place where the mirror is deposited 
by keeping the cotton thread wet (see sketch). 

By means of the apparatus described Lockemann has detected even o.oooi mg. 
of arsenic distinctly. 

Moist air gradually oxidizes the arsenic mirror, but in an absolutely dry at- 
mosphere even when exposed to light there is no change. In a closed tube con- 
taining a little phosphorus pentoxide arsenic mirrors may be kept unchanged even 
for months. 

Glass wool, or cotton, noticeably decomposes arsine. The decomposition of 
arsine in aqueous solution is also hastened by the presence of fine filamentaiy 
bodies. This reaction is probably catalytic in character. 

Electrolytic Estimation of Minute Quantities of Arsenic 

(C. Mai and H. Hurt») 

By this method minute amounts of arsenic (fractions of a 
milligram) are separated quantitatively at the cathode from an 
arsenical electrolyte as arsine. The latter then reacts quanti- 
tatively with silver nitrate as follows; 

AsH, + 3H1O + 6AgN0, = H,AsO, + 6HN0j + 6Ag. 

The advantages of the electrolytic detection of arsenic are 
first the avoidance of traces of arsenic that sometimes come 
from zinc in the Marsh test and second that destruction of or- 
ganic matter is often unnecessary. T. E. Thorpe* has shown the 
latter to be the case in the examination of beer worts and malt 
extracts for arsenic. To reduce arsenic add and its salts, a few 
drops of zinc sulphate solution should be added to the sulphuric 
acid acting as the electrolyte. The cathode is said to have a 
higher tension and the hydrogen to be very active. 

Apparatus and Procedure. — The apparatus used by Mai and 
Hurt is shown in Fig. 20. 

A is the reduction tube and B a bulb tube with 5-6 bulbs con- 

^ Zeitschrift fUr Untersuchung der Nahrungs- und Genussmittel 9, 193 (1905) 
and also Pharmazeutische Zeitung, 1905. 

• Proceedings of the Chemical Society 19, 183 (1903). 



SPECIAL QUALITATIVZ AND QUANTITATIVE METHODS 231 

taining 0.01 n-silvei nitrate solution. A and B are connected 
by a small tube g containing pieces of pumice stone saturated 
with an alkaline lead solution, or glass wool, to retain any traces 
of hydrogen sidphide. Anode a and cathode e are lead strips 
about 1-2 mm. thick. Their upper ends about 5 mm. thick 
are luted into glass tubes b which pass through the stopper of the 
U-tube and are tight. The dropping funnel d holds about 25 cc. 




i.**^ 



Flc. 30. — Apparatus for the Electrolytic Estimation of Arsenic. 

and its capillary end dips about 2 cm. into the solution to be 
electrolyzed. Tube c for the escape of oxygen from the anode 
chamber contains a little water. 

Fill U-tube A up to the mark with 12 per cent, arsenic-free 
sulphuric acid and bulb tube B with 10 cc. of o.oi n-silver nitrate 
solurion. Turn on the current and keep at 2-3 amperes. If 
the silver nitrate solution remains unchanged after hydrogen 
has been running i hour, the lead cathode and sulphuric add 
are arsenic-free. Without stopping the current, introduce from 
the droppii^ funnel the solution to be tested for arsenic, the 
quantity of which should not be more than 10 cc. Add this 
solution as slowly as possible and wash the last traces in with a 



232 DETECTION OP POISONS 

little water. If the solution contains arsenic^ or arsenic add, 

the silver nitrate solution will become dark in a few minutes and 

the reaction will be at an end in 3 hours. Pour the contents of 

the bulb tube through a small asbestos filter, wash with 3-4 cc 

of water and titrate excess of o.oi n-silver nitrate with o.oi 

n-potassium sulphocyanate according to Volhard's method. 

CalculatioiL — The reaction above shows that 6 molecules of 

silver nitrate correspond to i atom of arsenic (■* 7S). Therefore 

I gram-molecule of silver nitrate =1/6 gram-atom of arsenic = 

75 

^ = 12.5 grams of arsenic and 1000 cc. of o.oi n-silver nitrate 

= 0.125 gram of arsenic. 

Notes. — Electrodes of platinum (foil or gauze) cannot be used in the electro- 
lytic separation of arsenic as arsine, because either soild arsine or elementary 
arsenic is formed. Mai and Hurt also found that gold, silver and tin cathodes 
gave unsatisfactory results and carbon electrodes were not much better. Pure 
lead alone meets all the requirements as a material for the electrodes. Oxygen 
compounds of arsenic are quickly and completely reduced to gaseous arsine only 
upon cathodes of absolutely pure lead. The attachment of a platinum wire to a 
lead electrode was sufficient to cause incomplete reduction of arsenic compounds. 
For this reason the electrodes consist of one piece of lead^ without soldering on 
wire of another metal. The best electrolyte is 12 per cent, sidphuric add. A 
stronger add easily causes the formation of hydrogen sidphide and a weaker add 
has the disadvantage of lower conductivity and lower spedfic gravity. The 
electrolyte should be spedfically heavier than the solution to be tested to keep 
the latter from passing at once to the bottom of the reduction tube. 

Mai and Hurt found the following amounts of As: 

As found 
0.223 mg. 
0.099 03g. 
0.105 nig- 

For qualitative tests the bulb tube may be replaced by the 
drying and ignition tubes of the Marsh apparatus. 

According to Mai and Hurt the statements of Thorpe and 
Trotmann, that every solution can be electrolyzed without pre- 
viously destroying organic matter, do not always hold. In the 
examination of beer containing arsenic the results were fairly 
satisfactory, but in the case of urine the results were far too high. 

^ Kahlbaum's purest lead. 





As taken 


AsaO, 


0.25 mg 


AstOs 


o.io mg, 


AssOs 


o.io mg 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 233 



Quantitathre EstiniatiQn of Arsenic and Antimony by the Gntzeit Method 

Using a special apparatus and paper sensitized with mercuric 
chloride, Sanger and Black^ have found that the Gutzeit test 
can be employed to determine small amounts of arsenic quanti- 
tatively. The process is very simple and requires only a short 
time for completion. Sanger and 
Riegd* have extended this miethod 
to the estimation of antimony. 

Sensitized Paper. — Paper strips' 
uniformly 4 mm. wide are sensitized 
by being soaked in 5 per cent, solu- 
tion of recrystallized mercuric chlor- 
ide. These are dried, cut into 7 cm. 
lengths and protected from light and 
moisture in a stoppered bottle con- 
taining calcium chloride, or soda 
lime, covered with cotton. 

Apparatus. — ^A 30 cc. bottle (Fig. 
21) for the reduction is closed by a 
glass stopper provided both with a 
thistle tube, constricted to 2 mm. at 
the end and extending nearly to the 
bottom of the bottle, and with an 
exit tube widened to about 15 mm. 
just above the stopper. Connected 
with this exit tube by a ground joint and at a right angle is a 
tube exactly 4 mm. inside diameter and approximately 9 cm. in 
length from the bend. 

Procedure. — Place 3 grams of uniformly granulated zinc^ in 

* Proceedings of the American Academy of Arts and Sciences 43, 297-324 
(1907). 

* Ibid., 45, 21-27 (1909). 

* A cold pressed paper made by Whatman has been found to give the best 
results. 

* This all glass apparatus, suggested by Mr. W. A. Boughton, is now in use in the 
Harvard laboratory and is a modification of Sanger's original apparatus. 

* Bertha spelter from the New Jersey Zinc Company, New York, has been 
proved free from arsenic. 




Fig. 21. — Apparatus for the 
Quantitative Gutzeit Method.* 



234 DETEcnox of poisons 

the bottle and a strip of sensitized paper in the4mni. dcpositkai 
tube. In estimating arsenic place in the enlargement of tiie 
exit tube a loose plug of clean absorbent cotton that has been 
kept over sulphuric add; an hour's prdiminary run is necessary 
to moisten the cotton partially. In the case of antimony sub- 
stitute for cotton a disc of filter paper that has been mobtened 
with normal lead acetate, dried and kept in a wdl stoppered 
bottle. Before inserting this disc moisten it with a drop of 
water. Xezt add 15 cc. of diluted hydrochloric add^ (i :6) 
and let the hydrogen run 10 minutes to make sure the reagents 
cause no stain. Then add the whole, or an aliquot part of the 
solution to be tested. Arsenic will produce a color on the paper 
in a few minutes which will reach a maximum within 30 minutes. 
Antimony produces no \'isible effect on the sensitized paper, 
unless the amount is above 70 mmgr. (=0.070 mg.) irtiena gray 
color may appear. If there is any color, another trial should be 
made with a smaller portion of solution. In the determination 
of arsenic a disc of lead acetate paper should be inserted beneath 
the cotton as a precaution against the possible formation of 
hydrogen sulphide. 

Standard Bands. — (a) Arsenic. Dissolve i gram of resub- 
limed arsenious oxide in a little arsenic-free sodium hydroxide, 
addify with sulphuric acid and make up to a liter with recently 
boiled water. Dilute 10 cc. of this solution (I) to a liter with 
freshly boiled water which gives a solution (II) containing 
o.oi mg. of arsenious oxide per cc. Using definite volumes 
of solution II, measured from a burette, prepare a series of color 
bands (Fig. 22), taking a fresh charge of zinc and add for each 
portion. The color ranges from lemon yellow through orange 
yellow to reddish brown. 

(6) Antimony. — Dissolve 2.3060 grams of pure, recrystallized 
tartar emetic in a liter of water. This solution (I) contains i.o 
mg. of antimonious oxide per cc. By dilution of (I) solutions 
containing o.oi mg. (II) and o.ooi mg. (Ill) are prepared and 
used in making sensitized bands. 

^ The Baker and Adamson Company of Easton, Pennsylvania, supply a very 
pure acid suitable for this test. 



film 



Fig, aiH.— Standard Arsenic Baiul- m \\-- ■ 




ml 


•mil 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 235 

These bands will eventyally fade but they may be preserved 
longer by being sealed in glass tubes in the bottom of which is 
phosphorus pentoxide covered with cotton. The color of the 
arsenic bands may be developed (i) by placing the band in hy- 
drochloric acid (i :i) for 2 minutes at a temperature not over 
60", washing thoroughly, drying and sealing as before; (2) by 
treating for a few minutes with ammonium hydroxide, which 
gives a dense, coal black color, washing, drying and sealing in a 
tube over quicklime. 

To develop the antimony band, let it stand in a test-tube 
covered with nortnal ammonium hydroxide 5 minutes. A 
black band is slowly developed. These bands may be protected 
as described, or placed between glass plates cemented together 
and bound with passepartout paper. 

The more dilute standard solutions must be freshly made up 
within a few hours of use. 

Notes. — Solutions should be as free u piusible from sulphur compounds 
yielding hydrogen sulphide; interfering oiganic matter; and metals retarding 
formation of arsint and stibine. The cotton in the exit tube should be replaced 
after 10-12 runs, and the lead acetate disc after each tun. H the solution 
<»ntains arseniate, reduce with lo ce. of arsenic-free sulphurous add and 
^pel the excess. 

The absolute delicacy of the method is set at o.ooooS mg, of arsenious oxide 
id 0.0005 ^S- of antimoniouB oxide. The practical delicacy, using a band 
4 mm. wide, is o.oot rag. of arsenious oxide and o.oo» or 0.003 mg. of anti- 
moniotis oxide. By tising, however, a band i mm. wide in a correspondingly 
naiTOW exit tube, a practical deUcacy of 0.0005 mg. of arsenious oxide and o.oot 
mg. of antimonious oxide is obtainable. In length of band and density of 
developed color, the effect of arsine on the sensitized paper is from 2-3 limes 
as great as that of stibine. The authors do not claim a greater accuracy for the 
method than within 10 per cent. 



Biological Detection of Arsenic by Peniciilium Brevicaule 

B. Gosio' was the first to show that certain moulds, grown 
upon media containing minute quantities of arsenic, produce 
■olatile arsenic compounds characterized by a garlic-like 
odor. Seven species of moulds were found to have this power. 

> "Azione di alcune mufle sui composti Gssi d'arsenico," Rivistn d'igiene h 
Bfuita pubtica, 1S92, 201. 



236 DETECTION OF POISONS 

Penicillium brevicaiile, however, which Gosio isolated from 
air, and which was first found upon decaying paper, possessed 
this property in the highest degree. Gosio states that we are 
justified in regarding Penicillium brevicaule as a living reagent 
for arsenic. Even o.ooooi gram of arsenic can be recognized 
with certainty by this biological test. The test is* so delicate 
that it should be of great value in tozicological analysis in the 
preliminary examination for arsenic. 

A. Maassen^ states that a temperature of 28 to 32^ is most 
favorable to the growth of the mould. Crumbs of wheaten 
bread were found to make an especially good culture-medium. 
When this material is used, a vigorous growth of mould is 
visible even in 48 hours. Sometimes a test for arsenic can be 
finished in a few hours, and always in 2 or 3 days. The char- 
acteristic garlic odor from weak, arsenical cultures can be dis- 
tinctly recognized even after several months. That these 
"arsenic moulds" do not produce gases having a garlic odor 
from sulphur, phosphorus, antimony, boron and bismuth com- 
pounds, is an important fact. But Penicillium brevicaule 
possesses in high degree the power of converting solid selenium 
and tellurium compounds into volatile substances having a pe- 
culiar odor. The odor, especially from tellurium cultures, is 
like that produced by arsenic cultures, namely, distinctly like 
garlic! The odor from selenium cultures, however, differs 
from that arising from arsenic cultures. It is more of a mer- 
captan odor. 

Biginelli^ found that the gases, generated from arsenic 
cultures by Penicillium brevicaule, are completely absorbed 
by mercuric diloride solution. Colorless crystals, having 
the composition (AsH(C2H6)2.2HgCl2), are formed. ' This 
is a double compound of mercuric chloride and diethyl arsine. 
This compound can easily be decomposed. It then diffuses 
an intense garlic odor. 

R. Abel and J. Buttenberg' state that a mould to be of 

^Arbeiten aiis dem Kaiserlichen Gesimdheitsamt, 1902, 478. 
*Chemisches Centralblatt (1900), II, 1067, and also (1900), II, iioo. 
'Zeitschrift fiir Hygiene, 32, 440 (1899). 






SPECIAL QUALITATIVE AND QUANTITATIVX METHODS 237 

use in the biological detection of arsenic must satisfy the fol- 
lowing conditions: "It must grow rapidly, and not generate 
any odors during growth, except the garlic odor produced from 
an arsenical medium. It must not be restricted as to culture 
medium. It must grow in presence of large, or very small 
quantities of arsenic. Finally, it must demonstrate its specific 
action in presence of metallic arsenic and all kinds of arsenical 
compounds." 

The best material for these experiments is white or Graham 
bread, either of which is a favorite culture medium for moulds. 
The crust is the only part of bread having a specific aromatic 
odor. When this has been removed, the crumbs may be said 
to be practically odorless. 

Procedure. — When the material examined is liquid, absorb 
it completely by adding bread crumbs, and scatter a small 
quantity of dry bread over the surface. Solid material should 
be finely ground, or cut into as small pieces as possible, and 
placed in not too small a flask. Add at least the same quantity 
of bread crumbs, thoroughly mix the two substances by shaking, 
and moisten the mass with a little water. Close the flask with 
a cotton plug, and sterilize in steam. Sterilization must kill all 
micro-organisms in the flask. Therefore, heat the flask in an 
autoclave lo to 30 minutes under a pressure of i to 1.5 atmos- 
pheres. There is no danger of volatizing arsenic during ster- 
ilization. Then inoculate the sterilized material when cold. 
Place in a flask a slice of potato, superficially coated with 
mould in the spore-forming stage, and agitate it with bouillon 
(peptone), salt solution or sterilized water, until it is finely 
disintegrated. Observe all necessary precautions, and add 
the mould, suspended in water, in sufiicicnt quantity to 
impregnate the entire surface of the material suspected of 
containing arsenic. There should not be more liquid, how- 
ever, than the culture medium will absorb. Too much mois- 
ture retards the growth of the mould. Finally, draw a tight 
rubber cap over the mouth of the flask and cotton plug. Flasks 
thus closed may stand in the room, but it is better to keep them 
at a higher temperature, for example, in an incubator at 37°, 



4 



238 DETECTION OF POISONS 

since these conditions are most favorable to the growth of 
the mould. As soon as a growth of mould is distinctly visible 
to the naked eye upon the medium, the first indication is given 
that a test of the culture for volatile arsenic compoimds may 
prove successful. In a very favorable case, this is possible in 
24 hours. There is always a luxuriant growth of mould in 48 
to 72 hours, so that a decision can be reached. If there is no 
odor, the flask is closed, and the test is repeated once or twice 
daily on the following days. 

Sulphuric, hydrochloric and other strong mineral adds prevent the growth 
of the mould. This preventive action may be overcome by neutralization 
with calcium carbonate, which may be present in excess without ill effect. 
Alkalies also interfere with the growth of the mould. They may be removed 
by neutralization with tartaric or citric add, either of which may be present in 
excess. The great advantage of the biological over the purdy chemical method 
lies in the fact that less time is required to get a result. The tedious and im- 
avoidable destruction of organic matter in the material is rendered unnecessary. 
Moreover, a number of tests for arsenic may be made at the same time. 

Abel and Buttenberg (loc. dt.) speak as follows, regarding this method: 
''The biological method of detecting arsenic has so many advantages, that it 
deserves to be recommended for the most varied purposes. Its application is 
very general, and the method of procedure b simple. The culture of the mould 
can be kept a long time, even a year or more, without being revived. The 
test is very delicate, and the odor is readily recognized. The generation of the 
odor, in the case of cultures containing only 0.000 1 gram of arsenic, can be 
demonstrated for a week." 

Besides bdng practically unlimited in application, the biological method is 
extraordinarily delicate. In this respect, it exceeds the best known chemical 
methods for detecting arsenic. It is, for example, considerably more delicate 
than Bettendorff's test, and it might equal in delicacy the Marsh and Gutzeit 
tests. 

Detection of Arsenic in Organic Arsenic Compounds 
Cacodylic Add, Arrhenal, Atozyl^ 

The ordinary reagents usually fail to show arsenic in an or- 
ganic arsenic compound dissolved in water. Several of these 
compounds persistently resist the most powerful oxidizing and 
reducing agents. 

Cacodylic Acid, (CH8)aAsO-OH, and its salts have been used 
of late as drugs. A 2 per cent, solution of sodium cacodylate, 

^ C. £. Carlson, Zeitschrift fiir physiologische Chemie 49, 410 (1906). 



I 



I 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 239 

(CH3)sAsO-ONa.3HiO, conducts the electric current very feebly 
but no arsine appears at the cathode. Bettendorff's reagent 
(stannous chloride-hydrochloric acid) does not cause separa- 
tion of arsenic from cacodylic acid even after evaporation with 
hydrochloric acid and potassium chlorate. If heated with 
stannous chloride-hydrochloric acid, cacodylic acid is reduced 
to the foul smelling cacodylic oxide, [(CH3)jA5]iO, recognized 
by its odor. Distillation of sodium cacodylate by Schneider's 
method with the strongest hydrochloric add gives no arsenic 
trichloride in the distillate. The arsenic changes to another 
form, not precipitable by hydrogen sulphide. Evaporation of 
the distillate upon the water bath with nitric acid leaves solid, 
non-volatile cacodylic acid in which arsenic may be detected by 
reduction with sodium carbonate-potassium cyanide mixture. 
Even fuming nitric acid does not oxidize cacodylic acid to ar- 
senious or arsenic acid. 

Arrhenal, Sodium Methyl-Arseniate, (CHB)AsO(ONa)j.5H20, 
forms white crystals very soluble in water. Possibly owing to 
partial hydrolysis, an aqueous solution of this compound is 
alkaline and conducts the electric current feebly. Only traces 
of arsine appear at the cathode after electrolysis in presence of a 
good conductor. In arrhenal the arsenic is not held as strongly 
as in the cacodyl compounds. Hydrogen sulphide precipitates 
yellow arsenic trisulphide. Distillation with strong hydro- 
chloric acid gives arsenic trichloride in the distillate. Betten- 
dorff's reagent gives a red-brown precipitate, if considerable 
arrhenal is present. 

Ato^l, the Anihde of Metarsenic Acid, AsOj.NH.CsHs, 
forms white, odorless crystals readily soluble in water and hav- 
ing a faint, saline taste. As compared with cacodylic acid, 
arsenic in atoxyl is less firmly bound. Electrolysis gives arsine 
abundantly at the cathode. Hydrogen sulphide precipitates 
sulphide of arsenic. Arsenic trichloride passes over, upon dis- 
tillation with concentrated hydrochloric acid. Bettendorff's 
reagent gives a lemon yellow precipitate. 

Urine. — In suspected arsenic poisuniog first examine the unne. since arsenic 
!• very slowly cKminnied by this channel. Carlson in experiments upon him- 



240 DETECTION OF POISONS 

self was able to detect arsenic direct in the urine by tlie electrolytic method and 
also by the Gutzeit and Marsh tests. He took lo drops of Fowler's solution^ 
daily. Five days after the last dose Carlson could still get a distinct test for 
arsenic in concentrated urine. The urine was not wholly free from arsenic until 
14 days had passed. He then experimented with sodium cacodylate, taking 
daily 20 drops of a i per cent, solution. He could not detect a trace of arsenic 
in the urine by the electrolytic method. Therefore the salt of cacod^c add 
had passed through the organism unaltered. But cacodylic add can be de- 
tected easily in the urine, upon treating the latter with hypophoq>hon>us add 
(sp. gr. 1. 1 5).' Cacodylic oxide b formed and can be recognized by its odor. 
Sometimes the mixture must stand several hours in a dosed test-tube. Arrhenal, 
in daily doses of about 30 drops of i per cent, solution, behaved like the cacodyl 
compound. Arsenic could not be detected in the urine by electrolysis. Con- 
sequently neither arsenious nor arsenic add had been formed within the organism. 
Hypophosphorous add immediatdy predpitated arsenic from arrhenal and gave 
the cacodyl odor. 

To detect cacodylic add in urine, phosphorous add, as well as zinc or tin and 
hydrochloric add, may be used instead of hypophosphorous add. Frequently 
it is advisable to oxidize most of the organic matter in the urine beforehand. 
Boil 25 cc. of urine with 25 cc. of water, 5 per cent, potassium permanganate 
solution and 10 cc. of 25 per cent, sodium hydroxide solution, imtil the filtrate 
is odorless and nearly colorless. Excess of hydrochloric add (sp. gr. 1.19) and 
zinc filings, added to this filtrate, produce with heat the odor of cacodyl, if the 
urine contains cacodylic add. Arsenic from atoxyl can be isolated at the cathode 
in the form of arsine by electrolysis. Therefore arsenic can be detected in the 
urine electrolytically after administration of atoxyl. 

Quantitative Estimation of Minute Amounts of Arsenic 

(Karl Th. M«mer») 

This method is said to be useful in estimating arsenic quantitativdy in various 
kinds of fabrics and in urine in cases of poisoning. It is a titration method de- 
vised especially for quantities of arsenic not exceeding 0.5 mg. Arsenic 
is first predpitated as trisulphide with thioacetic add, CHi.CO.SH. Under the 
conditions arsenious as well as arsenic add is thus predpitated. In alkaline solu- 
tion potassium permanganate readily oxidizes arsenic trisulphide completely to 
arsenic add and sulphuric add: 

As2Sj + 14O = As,Oi + 3SO1. 

Potassium permanganate solution, added to an alkaline solution of arsenic 
trisulphide, immediately loses its color, bdng decomposed in the proportion of 

^ Fowler's solution contains i per cent of AS2O9 as potassium arsenite. 

' Instead of free hypophosphorous add, prepare Engel and Bernard's arsenic 
reagent (Comptes rend, de I'Acad. des sdences 122, 399), or J. Bougault's (I. 
Pharm. Chim. (6), 15, 527). Dissolve 20 grams of sodium hypophosphite in 
20 cc. of water and add 200 cc. of hydrochloric acid (sp. gr. 1.17). Filter through 
a cotton plug to remove NaCl and use the filtrate. 

' Zeitschrift fiir analytische Chemie 41, 397 (1902). 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 241 

9 molecules to i molecule of Arsenic tdsulphide.^ Since 2 molecules of potassium 
pennanganate in sulphuric add solution yield 5 atoms of oxygen for oxidation, 
9 molecules according to the proportion 

2:5 — 9:x (x — 22.5) 

should give 22.5 atoms of oxygen. But according to the reaction above, only 
Z4 atoms of oxygen are used to oxidize i molecule of arsenic trisulphide, whereas 
the remaining 8.5 atoms are stored up in the precipitate as hydrated manganese 
dioxide (MnOt.HtO). But if the reaction mixture is heated with oxalic add in 
presence of dilute sulphuric add, these oxygen atoms become active: 



(Mn; 



SO4 



;0 jO) 


► CO! OH 

+ 1 i 


iH,j 


COOiH 



MnSOi + H,0 + 2CO1 + H,0. 



Since 2 molecules of KMnOi yidd 5 atoms of oxygen and since 14 atoms of 
oxygen are necessary for i molecule of AsaSs, according to the following pro- 
portion 

Atoms : Mols.KMnOi 

5 : 2 -14 :x (x-s.6) 

5.6 molecules of potassium permanganate are required for i niolecule of AstSi 
(» 214), or 2 atoms of arsenic (» 150). 
1000 cc. of o.oi n-potassium permanganate (s 0.3162 gram KMnO^ contain 

in solution — — — — = 0.002 gram-molecule of KMnOi which according 

10 X 10 X 10 ° 

to the proportion 

Gram-mols.KMnOi : Grm. As 

5.6 : 150 = 0.002 : X (x — 0.0536) 

represents 0.0536 gram of arsenic. Hence 1000 cc. of o.oi n-potassium per- 
manganate solution correspond to 0.0536 gram of arsenic. 

Procedure. — Dissolve arsenic trisulphide in 0.5 pei cent, potassium hydroxide 
solution' and run this solution into a small flask containing 25 cc. of o.oi n-potas- 
sium permanganate solution. Mix the contents and add 5 cc. of 5 per cent, sul- 
phuric add, as well as the quantity of o.oi n-oxalic add solution found necessary 
by spedal titration. Warm until the color is discharged and Anally titrate with 
o.oi n-potassium permanganate solution. 

Preliminary Titration. — Add the same quantity of 0.5 per cent, potassium 
hydroxide solution used to dissolve arsenic trisulphide, as well as 5 cc. of 5 per 
cent, sulphuric add, to 25 cc. of o.oi n-potassium permanganate solution. Heat 
the mixture to boiling and add oxalic acid solution in slight excess so that the 
liquid becomes colorless. Titrate back with o.oi n-potassium permanganate. 

^ Since 2 mols. KMn04 give in alkaline solution 3 atoms of available oxygen 
(2SLMn04 = 2MnOs 4- 3O -f K2O), i mol. of AsaSi, according to the pro- 
portion: 3 : 2 = 14 : X (x = 9.33), requires not 9 mols. but more exactly 9.33 
mols. of KMnOi. 

* Ammonium hydroxide cannot be substituted for potassium or sodium 
hydroxide solution. 
16 



242 DETECTION OF POISONS 

This titration shows how much oxalic add solution, in conjunction with traces 
of reducing substances that may be present in the potassium hydroxide solution 
or sulphiuic add, is needed in a regular titration for the exact reduction of 25 
c.c. of o.oi n-potassiimi permanganate. 

Example. — Suppose that 25.5 cc. of oxalic add solution were required to de- 
colorize the boiling liquid. Titration required 0.3 cc. of o.oi n-potaarium per- 
manganate solution. Therefore 25 + 0.3 » 25.3 cc. of o.oi n-potassium per- 
manganate correspond to 25.5 cc. of oxalic add solution and 25^ cc. of the former 
correspond to 25.2 cc. of the latter solution. Consequently in a regular titration 
25.2 cc. of .0.01 n-oxalic add solution must be used. 

The amount of potassium permanganate in 25 cc. of o.oi normal solution is 
sufficient for all amounts of arsenic up to 0.5 mg. Mdmer's method of 
determining arsenic gives very reliable residts if arsenic is in the form of the 
trisulphide and free from every other substance soluble in 0.5 percent, potassiimi 
hydroxide solution and capable of reducing permanganate. 

Using the strongest hydrochloric add, M6mer first distils arsenic as arsenic 
trichloride by the Schneider-Fife method. About 200 sq. cm. of carpet, 100 sq. 
cm. of other woven and paper materials and 15 grams of sealing wax, stearine 
or wax candles and dried apples were used for each determination of arsenic by 
this method. According to M5mer, the distillate from such materials by the 
Schneider-Fife method always contains organic matter, even when caught in 
dilute nitric add. To remove this organic matter before predpitating arsenic 
with thio-acetic add, collect the distillate in a receiver containing dilute nitric 
acid and evaporate to dryness in a porcelain dish. Add to the small residue in the 
dish upon the water-bath successively 2 cc. of potassium hydroxide solution 
(0.5 per cent. KOH) heating i minute, then 2 cc. of potassium permanganate 
solution (5 per cent. KMnOO heating about 3 minutes, and finally i cc. of 
tartaric acid solution (20 per cent. Hj.CiHiOe)^ heating until the color is dis- 
charged. Filter into a porcelain dish, wash the filter with a little water and set 
the dish upon a boiling water-bath. Add after i minute i cc. of thio-acetic add 
(S per cent. CHj.COSH)* and warm the mixture 3 minutes. Arsenic is predpi- 
tated as arsenic trisulphide. After cooling for 5 minutes, collect the predpitate 
upon a filter and wash first 5 times with 2 cc. portions of 0.5 per cent, siilphuric 
add and then 3 times with 2 cc. portions of water. Place under the funnel a 
small flask containing 25 cc. of o.oi n-potassium permanganate solution and pour 
over the filter 3 portions of 0.5 per cent, potassium hydroxide solution, using 2 cc. 
each time. The alkaline solution of arsenic trisulphide thus drops directly into 
the permanganate solution. Otherwise, proceed as described. Subtract 0.3 cc. 
of o.oi n-permanganate solution from the volume of this solution used. This 

* Tartaric add readily dissolves the precipitate of manganese peroxide. To 
reduce the latter, Morner used oxalic and lactic adds, sodium sidphite and also 
thio-acetic add. But tartaric acid proved to be better than any of these 
substances. 

* Prepare thio-acetic acid solution by shaking 5 cc. of thio-acetic add with 
100 cc. of water. Filter and keep this solution in a dark flask. This solution 
gradually decomposes with evolution of hydrogen sulphide: 

CHj.COSH 4- H,0 = CH,.COOH + H,S. 



H spi 



I 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 243 

correction is necessary because even the finer qualities of filter paper conlus 
traces of substances which dissolve in 0.5 per cent, potassium hydroxide solu- 
tion and reduce permanganate.' 

Note. — The procedure described separates arsenic triiulpbide (rom every other 
substance soluble ia 0.5 per cent, potassium hydroxide solution and capable 
of reducing potassium permanganate. This method ia accurate to 0.01 mg, 
of arsenic. 

Detection of Salicylic Add in Foods and Beverages 

Wine.^ — Place 50 cc. of wine in a cylindrical separating funnel 
with 50 cc. of a mixture of equal parts of ether and petroleum 
ether. Shake frequently, taking care not to form an emulsion 
but yet to mix the liquids thoroughly. Remove the ether- 
petroleum ether layer, pour through a dry filter, evaporate 
upon the water-bath and add a few drops of ferric chloride solu- 
tion to the residue which becomes red-violet if salicylic acid is 
present. But if the color is black or dark brown, add a few 
drops of hydrochloric add, dissolve in water, extract with ether- 
petroleum ether and proceed with the extract as just described. 

Meat and Meat Products.' — For experimental purposes add 
about COT gram of salicylic acid to some chopped meat. Ex- 
tract the finely divided material with 50 per cent, alcohol and add 
some milk of lime to the filtered alcoholic solution. Evaporate 
to dryness upon the water-bath and stir the residue with a slight 
excess of dilute sulphuric acid. Shake with ether without fil- 
tering, pass the ether extract through a dry filter and evaporate. 
Dissolve the residue in hot water and test the filtered solution 
with very dilute ferric chloride solution for salicylic acid. 

Milk.* — Mix ICO cc. of milk with 100 cc. of water at 60". 
Precipitate with acetic acid and mercuric nitrate solution, using 
8 drops of each, shake and filter. Extract the filtrate with 50 

' After passing through the entire process in several blank experiments, 
MOrner never obtained higher results for permanganate used. Consequently 
the method of washing described completely removes tartaric and thio-acetic 

* "Official Directions tor the Chemical Examination of Wine" of June Jjlh, 
1896, (German.) 

' "Agreements in regard to uniformity in inspecting and testing foods, housed 
hold supplies and other articles used in the German Empire '' Heft 1, 36. 

* Method of Ch. Girard, Zeitschrift fUr analytische Chemie 11,177 (i8Sj) and 
the above "Agreements" Heft I, 61. 



244 



DETECTION OF POISONS 



cc. of ether, evaporate the ether, dissolve the residue in 5 cc. 
of hot water and test the filtered solution for salicylic add with 
dilute ferric chloride solution (sp. gr. i. 005-1 .010). 

Maltol 

Maltol, CeHiOs/ is formed in the preparation of caramel from malt, possibly 
from maltose or isomaltose. Ether or chloroform extracts this substance from 
the condensed vapors given off during caramelization and also from beer-wort. 
Maltol crystallizes in monoclinic prisms and plates from a cM. saturated solution 
in 50 per cent, alcohol (Osann). Chloroform gives denser crystals. Tliis sub- 
stance dissolves with difficulty in cold water or benzene; more readily in hot water, 
alcohol, ether or chloroform; and is insoluble in petroleum ether. It dissolves 
in caustic alkaline solutions but is repredpitated by carbon dioxide. Maltol 
sublimes in shining leaflets and is volatile with water vapor. It reduces silver 
solution in the cold and FeUing's solution with heat. An aqueous maltol solu- 
tion resembles salicylic add in becoming intense violet with ferric chloride solu- 
tion, but differs from carbolic and salicylic adds in not turning red with Millon's 
reagent. 

Maltol, shaken with benzoyl chloride and sodium hydroxide solution, gives 
a mono-benzoyl derivative and consequently must contain one hydroxyl 
group. 

Aqueous Chloral Hydrate Solution as a Solvent for Alkaloids, Glucosides 
and Bitter Principles and Its Use in Toxicological Analysis 

Richard Mauch 

(Communication from Professor £. Scbaer's Institute, Strassburg) 

One part of water at 17.5° dissolves 4 parts of chloral hydrate, forming a very 
mobile solution which is easily filtered and capable of being kept for a long time 
without decomposition. This 80 per cent, solution of chloral hydrate easily 
dissolves relatively large quantities of alkaloids and glucosides without altering 
them chemically. At 17.5° one part of each of the following substances 
requires for solution the number of parts of solvent stated in the table: 

Chloral Hydrate 

Solution 

(80%) Water Ether Chloroform 

Atropine 5 600 50 3.5 

Quinine 6 2000 freely soluble 2 

Cocaine 5 700 freely soluble freely soluble 

Morphine 5 5000 1250 100 

Santonin 4 5000 125 4 

Strychnine 6.5 6600 1300 6 

Brucine 6.5 .... .... .... 

Veratrine 7.5 .... .... 

^J. Brand, Berichte der Deutschen chemischen Gesellschaft 27,806 (1894), 
H. Kiliani and M. Bazlen, Ibidem 27, 31 15 (1894). 





SPECIAL QCAlITATIVi: AND QUANTITATIVE METHODS 

Caffdne b the only aUsloid which (orms with chloral hydrate a molecular 
compound soluble in water. If a chloral hydrate solution of an alkaloid, which 
haa been freshly prepared in the cold, is diluted with considerable 
unchanged alkaloid ia precipitated almost quantitatively, for instance, morphine, 
strychnine and qiunine. Substances like picrotoxin, santonin and acetanilide 
behave simikrly. Bui when such solutions stand tor a long time at ordinary 
temperatures, or are heated for i-i hours, chloral hydrate is decomposed by 
the vegetable base into chloroform and formic acid. Since the atkatoidol salts 
of formic acid ate soluble in water, dilution with this solvent does not precipitate 
the alkaloids. R. Mauch has shown clearly that atropine, brucinc, quinine, 
cocaine, morphine, narcotine, strychnine and veratrine behave as just described. 
In the tests ordinarily made with the ether or chloroform residue, R. Mauch 
recommends dissolving the residue in So or 60 per cent, chloral hydrate solution. 
The "chloral solution" should prove of great value in color tests which depend 
upon the use of pure sulphuric acid or sulphuric acid containing iron or molybdic 
acid. These solutions contain so httle water that it cannot modify the action 
of sulphuric acid upon the substance in solution. Such a "chloral solution" is 
also well adapted for zone tests. An aqueous solution forms an upper layer with 
the "chloral solution," and the latter forms an upper layer with concentrated 
sulphuric acid. 

Specific gravity of 80 per cent, chloral hydrate solution " 1.514. 

Specific gravity of 6oper cent, chloral hydrate solution -^ 1-35J5. 

In the tests ordinarily performed in test-tubes, it is best to use small tube 
(6 or 7 cm. high; t cm. in diameter) holding 6 cc. They should not be made of too 
thin glass. The chloral solution cannot be used in detecting picrotoxin, because - 
chloral hydrate itself produces the same reduction changes caused by picrotoxin. 
The same is true of the test for strychnine, where sulphuric add and potassium 
dichromate, or any other oxidizing agent, are used. Coniine and nicotine also 
belong to the class of alkaloids which cannot be detected in chloral hydrate solu- 
tion. Concentrated chloral hydrate solutions cannot be used directly in making 
teats with general alkaloidal reagents, because precipitates do not appear until 
the solutions have been diluted with 6-S volumes of very dilute hydrochloric or 
sulphuric aad. 

In using the "chloral hydrate method" in toxicological analysis, the etlwr, 
chloroform or amyl alcohol extract should be evaporated with gentle heat upon » 
watch glass of medium si2e (about S cm. diameter) and not too flat. Add to the 
residue, depending upon the quantity, about 3 cc. of 75 percent, chloral hydrate 
solution. Cover the glass and let it stand for some time. Occasionally tilt 
the glass and bring the solution thoroughly in contact with the residue. Pass the 
solution through a very small filter, if necessary, and wash both watcii glass and 
filter with a few drops of pure chloral hydrate solution. Use this chloral hydrate 
solution for the individual tests. In testing for strychnine, evaporate a part of 
the chloral solution to dryness upon the water-bath. Warm, until the residue 
does not smell of chloral, and then test for strychnine with sulphuric add and 
potassium dichromate. 

To recover from the chloral hydrate solution most of the alkaloids and sub- 
, stances like picrotoxin, acetanilide and phenacetine, add excess of sodium hy- 



]Mtl DETECTION OF POISONS 

vli\u(M«^ «i4utUm and extract thoroughly with a little cUorofonn. The ''dilond 
Kyvb^^i^ m<>thiHl" li conducive to veiy neat work and this is a great advantage. 
'X%^ uiHk uf nv«»tallic utensils like knives and spatulas is entirdy unneoeasaiy. 

ESTniATION OF ALKALOIDS 

1. Picrolonate Method of H. Matdies^ 

K norr* guve the name picrolonic add to i-p-mtrophenyl-3- 
mcthyl-4-isonitro-5-pyrazolone. This compound is formed by 
Iho Uition of nitric acid upon methyl-phenyl-pyrazolone. 
IMorolonlo acid resembles picric add in its properties and is 
iharai terUiHl by forming crystalline salts with many organic 
baskes^i us the alkaloids. As a rule these salts dissolve with 
iUtVu uU y und are yellow or red. Heat causes their decomposi- 
ti\U\. IMorolonic add is frequently of service in characterizing 
bii^O!i. Hydrivhloric add predpitates this compound from a 
:ii\\l\Uism \N| its so^lium salt as a yellow, mealy powder, melting 
\\hcu r^ixidly heated at about i^S^, becoming dark in color and 
uuvloTjivxiivs decomposition with rapid e\x»lution of gas. Knorr 
livst ^.-^ve piorv\lonic acid formula I but now formula IP is 
jweierrwi: 



X 0-OH 




\t M^^ttheo^ h^ e^umar^d t:::any alkak^ds qoantitaLtivdy by 
u\c4\\> xM \Nktv>!k>i\K ^id. Cvx,<ct ;be pffvdpttated alkalnidal 
^\w\\\kviutc i^^ A >axrij4!j<c Gvxvii cr;x£Sie. w^fesh. dry and wci^ 
V:nUuu;>s>;\ nNJ A,laX>kJs is ivtjsaSle by ^is zs^dicid. becanse die 



' Vx.xvX.s vV^- ;\N,^NK^<a ^<V^4u»s^>i» >*JMU&v'lit^t ^» 4^«. ^^terV 



SPEaAL QUALITATIVE AND QUANTITATIVE METHODS 247 

I SBtimatioa of Morphine, Codeine end Styptidne in Solutions, Tablets 
and Sugar Triturations 

Dissolve the weighed trituration or tablet in the smallest 
quantity of water possible and add picrolonic acid solution 
(about O.I n-solution in alcohol) in slight excess. The picro- 
lonate separates at once, or very soon, as yellow crystals or a 
crystalline meal. Cool 15-30 minutes in ice water and collect 
the precipitate in a weighed Gooch crucible. Wash with a 
little ice water, dry 30 minutes at 110° and weigh. Morphine 
picrolonate usually separates in 10-30 minutes. Cooling aids 
precipitation. 

Formula MdI. Wl. Deoompoiitlon-poiiit ' 
Morphine jMCToioDaie: CivHuNOi.CioHiNiOi. 545 300-310° 
"^Nldiie picrolonate: CnHnNOi-CioHsNiOi. 563 about 115' 
^Olamine picrolonate; CiiHuNOi.CioH.NtOt. 501 205-110° 



H'BeE 



Kotesj — Piftctice Analyses: Morphine powder: o.oi-o.os giam morphine hy- 
loride, C1TH11NO1.HCI.3H1O + 0.5 gram sugar. 0.3-0.5 gram codeine 
phosphate, Ci.H,iN0i,HiP0».2H,0 + 0,5 gram sugar. Stypticine tablets E. 
Merck. 

Do not use too dilute solutions of the alkaloids in these determinations and do 
not Wftsb the picrolonate precipitates with too much water. Dissolve the pow- 
dered morphine and sugar mixture in about 5-10 cc. of water. Matthes and 
Kammstedl in examining the morphine powder obtamcd the following results: 

Weights taken: a.019 morphine hydrochloride + 0.5 gram sugar. 
Besults obtained; 

,0273 gram morphine picrolonate = o.oiSj gram morphine hydro- 
gram morphine picrolonate = o.otS; gram morphine hy- 
drochloride. 

In a second experiment every 10 cc. of an aqueous solution contained 0.0104 
gram of morphine hydrochloride, 
Seeults obtmned: 

gram motphme picrolonate = o.oioi gram morphine hydro- 



_ Aceuiu t 






0.0146 gram morphine picrolonate = o.oogQ gram morphini 
chloride." 

The application of the picrolonate method to the estimation of hydrastine in 
hydrastis root and extract, of nux vomica alkaloids in nui vomica and ex tract and 
of pilocarpine in jaborandum leaves is described in Chapter VI (see pages 375, 
979 and 3S9. 

■ Professor Matthes has kindly stated that this picrolonate method gives less 
ifitisfactOTy results with smaller quantities of morphine (0.005 and less). 



248 DETECTION OF POISONS 

2. Estimation of Alkaloids by Means of Potasafaim Bisfimfhoos Iodide 

(H. ThomsO 

Dissolve the particular alkaloid in sulphuric add and pre- 
cipitate completely with potassium bismuthous iodide prepared 
as described by Kraut.^ Decompose the precipitate with a 
mixture of sodium carbonate and hydroxide, extract the free 
alkaloid with ether and weigh. By this method Thoms has 
recovered atropine, hyoscyamine, scopolamine, strychnine, 
quinine, caffeine and antipyrine from their potassium bis- 
muthous iodide precipitates unaltered and nearly quantitatively. 
He has also used this method with success in estimating quanti- 
tatively the alkaloids in belladonna extract. 

Procediire. — Dissolve the alkaloidal salt, or 2 grams of bella- 
donna extract, in 50 cc. of water. Add first 10 cc. of 10 per cent, 
sulphuric add, stir and predpitate with 5 cc. of potassium bis- 
muthous iodide solution. Collect the predpitate upon a dry 
filter and wash twice with 5 cc. portions of 10 per cent, sulphuric 
add. Transfer the thoroughly drained predpitate and paper 
to a wide-mouth extraction cylinder having a tight glass stopper. 
Add 0.3 gram of sodium sulphite, then 30 cc. of 15 per cent, so- 
dium hydroxide solution and shake. Add quickly 15 grams of 
sodium chloride and 100 cc. of ether. Shake frequently and let 
stand for 3 hours. The ether contains the alkaloid and settles 
well. Remove with a pipette 50 cc. of the ether solution (= 
half the solution of the alkaloidal salt, or i gram of belladonna 
extract) and titrate this ether solution in a flask with o.oi 
n-hydrochloric add, using iodeosine as indicator. 

After titrating the belladonna alkaloids, use in the calculation 
the equivalent weight of atropine-hyoscyamine, C17H28NO8 = 
289. 1000 cc. of 0.01 n-hydrochloric add correspond to 2.89 
grams of atropine-hyoscyamine. Atropine and hyoscyamine 
bdng isomeric, monadd bases, thdr formula wdght and equiva- 
lent weight are the same. 

^ Berichte der Deutschen phannazeutischen Gesellschaft 13, 240 (1903); 15, 85 
(1905); 16, 130 (1906) (D. Jonescu). 

* Annalen der Chemie und Pharmazie 210, 310 (1882). See "Preparation of 
Reagents," page 311. 



SPECIAL QUALITATIVE AND QDAKTITATIVE ItETHODS 

Quinine, caffeine and antipyrine were also recovered un- 
altered from potassium bismuthous iodide precipitates. After 
decomposition of the precipitates, they were obtained almost 
quantitatively but were estimated gravimetrically. 

Dissolve 2 grams of quinine^ in 50 cc. of water acidified with 
sulphuric acid and precipitate with potassium bismuthous iodide. 
Filter the precipitate with suction and wash with 5 per cent, 
sulphuric acid. Transfer precipitate and paper to an extraction 
cyhnder and shake thoroughly with a mixture of 20 grams of 
crj^tallized sodium carbonate and 40 cc. of 10 per cent, sodium 
hydroxide solution. The yellowish red precipitate gradually 
becomes white. Add 30 cc. of ether and shake well for 30 
minutes. Pipette off 25 cc. of the clear ether solution, evap- 
orate in a weighed glass dish, dry the residue at 100° and weigh. 
The weight of quinine was 0.9405 instead of i gram. 

Caffeine was estimated in the same way, except that this 
alkaloid was extracted with chloroform after decomposition of 
the potassium bismuthous iodide precipitate with alkaline 
hydroxide and carbonate. The weight of caffeine was 0.9546 
instead of i gram. 

The precipitate obtained by adding potassium bismuthous 
iodide to a solution of antipyrine in sulphuric acid (10 per cent. 
HbSO*) is not decomposed as easily as are those of quinine and 
caffeine. The precipitate from 2 grams of antipyrine must be 
shaken i hour with 20 grams of sodium carbonate and 60 cc. 
of 10 per cent, sodium hydroxide solution. Antipyrine must be 
extracted with chloroform. The weight of antipyrine was 
0.9273 instead of 1 gram. 

Holes. — Potassium bUmuUious iodide precipitates fixed and volatile 
alkaloids but not ammonium salts. If the estimation of volatile bases is unnec- 
essary, as in tbe eiamination of belladonna extract, evaporate tbe 50 cc. of ether 
extract (see above) upon the water-bath. ' Worm the residue and in a few min- 
utes the strong narcotic odor of volatile bases will disappear. Dissolve the 
residue in a little add-free alcohol and dilute with ether. Before using a flask 
for titrations carefully test it beforehand for alkalinity. If a positive test is 
obtained, alkalinity must be removed. An odor like iodoform, probably due to 
the action of sodium hypoidite upon the alkaloid, has been observed when sodi- 

' According to experiments of D. Jonescu (}oc. cil.). 



250 DETECTION OF POISONS 

um hydroxide solution acts upon potassium bismuthous iodide prediiitates. 
Addition of sodium sulphite may prevent this action. After addition <^ sodium 
chloride ether takes up the alkaloid more readily. But vigorous glu^lrtng is 
always needed to cause complete transfer of alkaloid to the ether. 

3. Estunation of Alkaloids by H. M. Gordm^ 

Gordin has f oimd that periodides of the alkaloids, whatever be their composi- 
tion, when precipitated from aqueous solution by iodo-potassium iodide in pres- 
ence of adds, always contain one equivalent of combined add for every molecule of 
monadd alkaloid. These periodides have»the general formula (Alkaloid, HI)inIii- 
Iodo-potassium iodide, added to a solution of a monadd alkaloid addified with 
hydrochloric add, first gives an alkaloid hydrochloride, changed by potassium 
iodide to alkaloid hydriodide and finally preapitated as insoluble periodide by tak- 
ing up iodine: 

(a) Alkaloid + HCl = Alkaloid.HCl, 

08) Alkaloid.HCl + KI = AlkaJoidHI -f KCl, 

(7) m(Alkaloid.HI) + In = (Alkaloid-HDn^I^ - Predpitate. 

In the predpitation of an alkaloid in add solution with iodo-potassium iodide, 
one equivalent of add goes with the precipitate and disappears from solution. 
In many cases potassium mercuric iodide may be substituted to advantage for 
iodo-potassium iodide. Gordin has found that the composition of the predpitate 
changes only as regards mercuric iodide and not as far as add is concerned, for in 
this case also the predpitate contains one equivalent of add for a monadd alkaloid. 
Use in the titration 0.05 n-hydrochloric add and 0.05 n-potassium hydroxide 
solution. Prepare a solution containing a weighed quantity of pure alkaloid, for 
example, pure morphine. Dissolve about 0.2 gram of chemically pure morphine, 
previously completely dehydrated at 120®, in 30 cc. of 0.05 n-hydrochloric add 
in a 100 cc. volumetric flask. Shake and add gradually iodo-potassium iodide 
to this solution, imtil predpitation ceases and the supernatant liquid is dark red. 
Dilute to the 100 cc. mark with water and shake vigorously until the liquid above 
the precipitate is entirely dear. Pass 50 cc. of solution through a dry filter, de- 
colorize the filtrate with a few drops of sodium thiosulphate solution and titrate 
excess of 0.05 n-hydrochloric add with 0.05 n-potassium hydroxide solution, using 
phenolphthalein as indicator. Caloilate from the result how many grams of 
morphine have been neutralized by i cc. of the add. Comparison of the equiva- 
lent weight of morphine with that of any other monadd alkaloid gives the corre- 
sponding factor to be used in the calculation. For example, Gordin found in his 
experiments that i cc. of approximately 0.05 n-hydrochloric add neutralized 
0.0137 gram of anhydrous morphine, CitHuNOj. The factor (x) for strychnine, 
C2iH22N20f (= 334), which is also a monacid base, is as follows: 

Morphine : Strychnine 

285 : 334 = 0.0137 : X (x = 0.0160) 

^ Berichte der Deutschen chemischen Gesellschaf 1 32, 2871 (1899). Archiv der 
Pharmazie 238, 335 (1900). Gordin and A. B. Prescott, Archiv der Pharmazie 
237, 380 (1899). 




Quantitative EstimatiDn of Stijclmine and Quinine Together 

(E. F. Harrison and D. Gait') 



SPECIAL QUALITATIVE AND QUANTITATIVE METHODS 251 

i tliat for the monacid base cocaine, Ci7HiiNDt(~ 303), according to the 

Motpli'n* - Cocalnn 

: 3°3 - O.OI37 : X (x = 0.0:46). 

If potassium mercuric iodide is used to precipitate an alkaloid, the method is 
the same as with iodo-potassium iodide except that there is no need of treating 
the so cc. of filtered solution with thiosulphate. 

IBerberine and colchicin cannot be estimated by Gordin's method. 
Occasionally a small amount of strj'chnine must be estimated in presence of a 
relatively large quantity of quinine, as in certain pharmaceutical preparations.* 
Separation of the tvo alkaloids is possible by means of Rocbelle salt. Quinine 
tartrate. {C,oH«N,0,),.C.H,Oc.3H,0, bang difQcultly soluble in water, forms 
a white crystalline precipitate, whereas strychnine tartrate remains in solution. 
Procedure. — Render the solution of tbc mixed alkaloids in about 40 cc. of 
water faintly add with sulphuric add. .^dd enough ammonia to cause a slight 
turbidity, then 15 grams of solid Rochelle salt and more ammonia, still leaving 
the liquid add to litmus paper. Heat for 15 minutes upon the water-bath and then 
set aside for a hours until entirely cold. Filter predpitated quinine tartrate by 
suction and wash with aqueous Rocbelle salt solution (15 grams of salt in 45 cc. 
of water) containing i-idropsof dilute sulphuric add. To determine strychnine, 
add sodium hydroxide solution to the combined filtrate and wash water ftora 
quinine tartrate until tlie reaction is alkaline. Extract 2-3 times with chloro- 
form, pour the chloroform extract through a dry filter and distil in a weighed 
flask to about 4 cc. Add 10 cc. of absolute alcohol and evaporate to dryness 
upon the water-bath. To remove quinine still adhering to the strychnine, ex- 
tract the dry residue j-3 limes with i cc. portions of ether,' dry at 100° and 
weigh. This residue consists of pure, quinine-free strychnine. 



■ Vandt 



ition of Toxicity of Chemical Compounds by Blood Haemolysis 
(A. J. J. Vandevelde') 
Vandevelde originally used li\Tng cells of a variety of Allium cepa (red Bruns- 
wick onion), the cell membrane of which is rich in anthocyan. The presence of 
this substance obviated the necessity of using a special coloring matter in 
determining plasmolytically the toiticity of alcohols, ethereal oils and other sub- 
stances.* Vandevelde has recently recommended determining the toxidty of 
chemical compounds by blood bimolyMS. using for this purpose defibrinated ox 

' Pharma*. Journ. (4) 17. i^s. 

' Compound Syrup of Hypophosphiles, U. S. P. 

• More ether dissolves a weighable quantity of strychnine. 

* Chemiker Zeitung ap, S^S ('9os)- 

> Bulletin de 1' Association Belgie de Chimie 17, ]jj. 



252 DETECTION OP POISONS 

blood (see pages a i6 and 222) i To establish the toxicity of different alcohols, the 
concentration at which haemolysis just ceases is determined. A solution, in which 
blood corpuscles are not hydrolyz^ after a definite time but are hydrolyzed upon 
addition of the slightest trace of the substance being examined, is a non-toxic solu- 
tion for blood corpuscles, called by Vandevelde a "critical solution." 
The estimation requires: 

1 . A solution of 0.9 per cent, sodium chloride in 50 per cent, alcohol by volume.^ 

2. An aqueous 0.9 per cent, sodium chloride solution. 

3. A suspension of 5 per cent, defibrinated ox blood in 0.9 per cent, aqueous 
sodium chloride solution. 

Experiments are made in test-tubes. Place first in each of several tubes a.5 cc 
of the mixtures (of different concentration) of alcoholic and aqueous sodium 
chloride solution and then 2.5 cc. of the suspended blood. The end point for 
the appearance of haemolysis was set at three hours. 

Vandevelde's experiments with ethyl alcohol gave the following results; 

^ - , , ,. Alcoholic con- 

^ - Cc. of alcohohc 

Cc. of sus- ., ^, , 
J J ti J NaCl solu- 
pended blood 
*^ tion 

2.5 2 . 20 

2.5 2. IS 

2.5 2.10 

2.5 2.05 

2.5 2.00 

2.5 1-95 

2.5 1.90 

Consequently the critical solution of ethyl alcohol is one containing 19.5 cc. of 
absolute alcohol (CtHeO) in 100 cc, or 15.489 grams of CsHiO in 100 cc. The 
specific gravity of absolute alcohol being 0.7943, 19.5 cc. weigh 19.5 X 0.7943 » 

15.489- 

According to Vandevelde's experiments addition of methyl alcohol diminished 
the toxicity of ethyl alcohol, whereas the higher alcohols were found to be more 
toxic than the latter. If the toxicity of 100 parts by weight of ethyl alcohol is 
taken as 100, then 47 parts by weight of isopropyl, 29 parts of isobutyl and 12.5 
parts of amyl alcohol are isotoxic with that quantity of ethyl alcohoL Using 
the plasmolytic method with onion cells, Vandevelde obtained the following results 
with the same series: 

100, 36.8, 21.2, 12.6. 

The hemolytic method is easily periormed in test-tubes and does not require 
the use of the microscope. The form of the tube, especially its diameter, is qtdte 
important in these experiments. The speed of haemolysis increases with the 
diameter of the tube. The quantity of the blood corpuscles is of slight influence 
except in narrow tubes and at the beginning of the reaction. Vandevelde applies 
the term ''critical coefficient" to the number giving the concentration of a sub- 
stance necessary to kill the cells. 

* The specific gravity of such an alcohol at 15® is 0.9348. 



Cc. of aqueous 


centration of 


After 3 


NaCl solution 


mixture in voL- 
per cent. 


hours 


0.30 


22.0 


Haemolysis 


0.3s 


21. S 


Hemolysis 


0.40 


21.0 


Haemolysis 


0.4s 


20.5 


Haemolysis 


0.50 


20.0 


Haemol3rsi8 


OSS 


19. S 


No haemolysis 


0.60 


19.0 


No haemolysis 



CHAPTER VI 

QUANTITATIVE ESTIMATION OF ALKALOIDS AND OTHER 

PRINCIPLES 

EBtiination of AUcaloids in Drugs and Phannaceutical Preparations 

(German Pharmacopoeia) 

Alkaloids are nitrogenous bases occurring in plants. The 
term "plant base" is synonymous with alkaloid. The plant 
families especially rich in alkaloids are berberidese, dnchonacese, 
papaveracese, solanacese and strychnaceae. Alkaloids as a rule 
are not uniformly distributed in all parts of plants. They 
occur most often in roots, fruits and seeds. If the plant is a 
tree, there is often more alkaloid in the bark than in other parts. 
The particular part of the plant usually contains only a few per 
cent, of alkaloid. Quinine bark is an exception, the quantity 
of alkaloid being 5-10 per cent, and sometimes more. Plants as 
a rule do not contain free alkaloids but their salts. They are 
combined not only with the mineral acids, sulphuric, hydro- 
chloric and possibly phosphoric, but with organic acids, as malic, 
aconitic, tannic, citric, quinic and meconic. Most free plant 
bases dissolve only slightly in water but readily in ether and 
chloroform. The German Pharmacopoeia prescribes a mixture 
of ether and chloroform for the extraction of free alkaloids. 
The finely powdered drug should first be treated with a solution 
of sodium hydroxide, ammonia or sodium carbonate to liberate 
alkaloids from their salts: 

C,oH,4N,0, 

C,H7(OH)4COOH + NaOH - CwH,4N,0» + C6HT(OH)4COONa + H,0. 

Ouinine quinate* Quinine Sodium quinate 

The ether-chloroform mixture removes not only alkaloids 
from the drug but varying amounts of other substances, as fat, 
resin, wax and pigments. To free the alkaloid from such im- 

^ Cinchona bark contains quinine in the form of this salt. 

253 



254 DETEcnox of pqisoss 



purities, shake the ether-chlonrfonn crtnct with a measured 
excess of o.i cm* o.oi n-hydrochl<mc add. The alkaloid passes 
into aqueous scdution as hydrochk>ride: 



But the impurities remain in the ether-cfakirofonn mixture. 
Finally, determine excess of hydrochk»ic add by titratioii with 
o.i or o.oi n-potassium hydroxide sohition, emi^oying usually 
iodeosine as indicator. Calculate the amount of alkalmd in the 
drug from the difference between the original quantity <rf add 
and the excess. 

The estimation of alkaloids in drugs and pharmaceutical 
preparations, according to directions given by the German 
Pharmacopceia, requires the following steps: 

1. Liberation of alkaloids from salts by means of stronger 
bases, as potassium and sodiimi hydroxides, ammonia and 
sodium carbonate. 

2. Extraction of free alkaloids with ether-chloroform mixture. 

3. Transference of alkaloids from ether-chloroform to aque- 
ous 0.1 or O.OI n-hydrochloric add solution. 

4. Determination of excess of hydrochloric add in an aliquot 
volume, usually 50 cc. of the hydrochloric add solution of the 
alkaloid diluted to 100 cc. by titration with o.i or o.oi n-potas- 
sium hydroxide solution. 

AlkakMds m Aconite Root 

Officinal aconite root is the root of Aconitum Xapellus col- 
lected at the end of flowering. Two alkaloids are present, 
namdy, aconitine, C»4H47XOii, and picraconitine, CjiBLnNOit 
(?), characterized by its verj- bitter taste. Both alkaloids are 
combined with aconitic add, 

CH.COOH 

C.COOH 

CH,.COOH 



QUANTITATIVE ESTIMATION OF ALKALOIDS 255 

Boiled with water, or alcoholic potassium hydroxide solution, 
aconitine yields a new base, aconine, benzoic and acetic acids :^ 

C,4H47NOii -f 2H,0 = C,»H4iN0t + C»H40» + C,H».COOH. 

Aconitine Aconine Acetic Benzoic acid 

acid 

Thiis reaction presents aconitine as acetyl-benzoyl-aconine, 

/COCH, 

Since aconitine has been shown to contain four methoxyl 
groups, the formula of this alkaloid may be written : 

/COCH, 
CHHr(0CH,)4N0»< 

Aconine, therefore, is C2iH27(OCH3)4(OH)2N03, and picra- 
conitine must be regarded as benzoyl-aconine, having the for- 
mula C2iH2i(OCH3)4(OH)N04(COC6H5). 



Estimation of Aconitine 

(German Pharmacopoeia) 

Place 12 grams of rather finely powdered aconite root dried at loo® in an 
Erlenmeyer flask and add 90 grams of ether and 30 grams of chloroform. Shake 
well and add 10 cc. of a mixture of 2 parts of sodium hydroxide solution and i 
part of water. Let the mixture stand 3 hours, shaking vigorously at frequent 
intervals. Then add 10 cc. of water, or enough to cause the powder to gather 
into balls after vigorous shaking and leave the supernatant ether-chloroform 
solution perfectly clear. After an hour pass 100 grams of the clear ether- 
chloroform solution through a dry filter kept well covered and receive the filtrate 
in a small flask. Distil about half the solvent and pour the remainder into a 
separating funnel. Wash the flask with three 5 cc. portions of a mixture of 3 
parts of ether and i part of chloroform, and thoroughly shake the combined 
solutions with 25 cc. of o.oi n-hydrochloric acid. When the liquids have 
separated perfectly clear, add enough ether to bring the ether-chloroform solu- 
tion to the surface. Pass the add solution through a small filter moistened with 
water and collect the filtrate in a 100 cc. flask. Make three more extractions 
of the ether-chloroform solution with 10 cc. portions of water, and pass these 
extracts through the same filter. Wash the latter with water and dilute the total 
solution with water to 100 cc. Place 50 cc. of this solution in a 200 cc. flask with 
50 cc. of water and enough ether to make a layer about i cm. thick. Add 5 
drops of iodeosine solution and run in, while shaking, enough o.oi n-potassium 
hydroxide solution to turn the aqueous solution pale red. 

* Freund and Beck, Berichte der Deutschen chemischen Gesellschaft 27, 
433, 720 (1894); 28, 192, 2537 (189s). 



256 DETECTION OF POISONS 

Calculation. — Dissolve the aconite alkaloids set free firom tlieir salts by 
sodium hydroxide solution in 120 grams of ether-chloroform. Weigh zoo grams 
of this solution (» alkaloids from 10 grams of aconite root). Dissolve the 
alkaloids with 25 cc. of o.oi n-hydrochloric add, bringing the vcdume to zoo cc. 
Determine excess of add in 50 cc of this solution (-^ alkaloids firom 5 grams d 
root). If, for example, this requires 8.5 cc of o.oz n-potassium hydroxide 
solution, then the alkaloids have combined with 12.5 — 8.5 -^ 4 cc of o.oz n-add. 
Since the equivalent weight of aconitine CiiHiTNOn » 645, 100 cc of o.oz 
n-hydrochloric add imite with 6.45 grams of aconitine. The proportion 

1000 : 6.45 ■* 4 : X (x » 0.0258) 

shows that 5 grams of aconite root contain 0.0258 gram of alkaloids, correspond- 
ing to 0.51 per cent. The German Pharmacopceia demands this quantity of 
aconitine in aconite root as a minimum. Using a different method, the United 
States Pharmacopoeia has the same limit. 



EstimatiQii of Cantharidin in Spanish Flies 

(German Pharmacopoeia) 

Place 25 grams of Spanish flies groimd mediumly flne in an Erlenmeyer flask 
and add 100 grams of chloroform and 2 cc. of hydrochloric add.^ Shake the 
mixture frequently during 24 hours. Then pour 52 grams of the chloroform solu- 
tion through a dry filter kept well covered, and collect the filtrate in a weighed 
flask holding 80-100 cc. Distil the chloroform, and add 5 cc. of petroleum ether 
to the residue. Stopper the flask, and let the mixture stand 12 hoiurs, with oc- 
casional agitation. Dry at 100^ and weigh a filter (5 cm. in diameter). Pass 
the liquid through this filter, having first moistened it with petroleum ether. 
Treat the undissolved residue twice with petroleum ether, each time using 10 cc. 
and shaking. Pass this solvent through the same filter, and disregard crystals 
adhering to the side of the flask. Dry the filter and flask, and wash both with 
a little water, containing a drop of ammonium carbonate solution to every 10 cc, 
until this solvent is only faintly yellow. Finally, wash once with 5 cc of water, 
and dry both flask and filter. Place filter and contents in the flask, and dry at 
100° to constant weight. The crystalline residue should weigh at least o.i gram. 

Notes. — Additional information about cantharidin is given 
on page 196. Spanish flies contain cantharidin partly free and 
partly as an .alkali salt of cantharidic acid (cantharidate). 
Hydrochloric acid sets cantharidic acid free and the latter thai 
passes at once into cantharidin, its internal anhydride (lactone). 
Consequently hydrochloric acid is essential to the detennination 
of that cantharidin present in Spanish flies as cantharidate. 
Chloroform not only dissolves cantharidin but fatty substances 

^ Specific gravity 1.124 = 25 per cent. HCl. 



QUANTITATIVE ESTIMATION OP ALKALOIDS 



257 



1 the flies. To isolate pure cantharidin from these impurities, 
istil the chloroform and let the residue stand for i2 hours hi the 
X)ld with petroleum benzene. Fat readily dissolves but can- 
ktharidin is as good as insoluble in this solvent. The German 
I Pharmacopceia finally directs weighing the cantharidin from 
, 12-5 grams of powdered Spanish flies. The quantity should be 
at lea.st o.i gram, corresponding to b.8 per cent, of cantharidin 
as a minimum. With sufficient care, white crystalline can- 
tharidin may be isolated from Spanish flies. 

Baudin obtained from good flies i.o6 per cent, of cantharidin, 
^of which 0,72 per cent, was free and 0.34 per cent, combined 
i cantharidate. Dieterich found only 0,3 per cent, of free 
ntharidin. 

Estimation of Cinchona Alkaloids 

(GermiLn Pharmacopceia) 

. In Cmchooa Bark. — To delETmine total alkaloids, pour go grams of Ether 
3 grams of chloroform upon ij gtams of finely ground cinchona bark, 
dried at 100° and placed in an Erlenmeyer flask. Add 10 cc. of sodium hydroxide 
solution. Shake vigorously at frequent inter\'als during 3 hours. Then add 10 
cc. of water, or enough to cause the powdered cinchona 10 gather into lumps 
after vigorous shaking, thus leaving the supernatant ether- chloroform solution 
perfectly clear. Let the ether- chloroform solution stand an hour, and then pau 
loa grams through a dry filter, kept well covered. Collect the filtrate in a Snsk. 
and distil half the solvent. Tour the remaining ether- chloroform solution into 
a separating funnel, and wash the flask thiee times with 5 ec. portions of a 
mixture of j parts of ether and t part of chloroform. Thoroughly extract the 
total ether- chloroform solution with 15 cc. of 0.1 n-hydrochloric acid. When the 
contents of the separating funnel are perfectly dear, add enough ether to bring 
the ether-chloroform solution 10 the surface. Pass the acid solution through a 
small filter moistened with water, and receive the filtrate in a 100 ec. flask. 
Make three more extractions of the ether-chloroform solution with 10 cc. porliona 
of water, and pass these extracts through the same 61lcr. Wash the filter with 
water, and bring the volume of the filtrate to 100 cc. Finally, measure 50 cc. 
of this solution with a pipette, and add freshly puepared biematoxylin solution, 
made by dissolving a small particle of this substance in 1 cc. of alcohol. Shake 
add enough o.i n-potassium hydroxide solution to give the mixture a yellowish 
r, which quickly changes after vigorous agilalion to bluish violet.' 

, Notes and Calculation.^ — Both quinine and quinitiine have 
i formula CjoH24N20a and dnchonine and cinchonidine the 

'*ThB German Pharmacopceia prescribes that not more than 4.3 cc. of 0.1 
m hydroxide should be required. 



258 DETECTION OF POISONS 

formula C19H22N2O. These are the most important 
in cinchona bark. They are present in all true cinchona barks 
as salts of quinic add, CnHijOe, andquino-tannicadcL Fuller 
information regarding the chemistry of quinine and dnchonine 
is given on page 114. 

Quinic acid is widespread in the vegetable kingdom. This 
monobasic, pentatomic add, having the formula, CtH7(OH)4- 
COOH, is a hexahydro-tetroxy-benzoic add. It crystallizes 
in large monoclinic prisms melting at 162^. As far as the 
chemical behavior of quinic add is concerned, either of the 
following formulas is possible: 

I. H OH II. H OH 

X ^ 

H,C CH.OH H,C CH.OH 

H,C CH.OH HO.HC CH, 

Y Y 

HO COOH HO COOH 

The formation of tetra-acetyl-quinic add, (CH^COO)!- 
C6H7COOH, and tetra-benzoyl-quinic add, (C6H5COO)4C6H7- 
COOH, shows that quinic acid contains four alcoholic hydroxyl 
groups. Addition of sodium hydroxide solution to cinchona 
powder sets the alkaloids free from their salts: 

CioH,4N,0, 

C,H7(OH)4COOH + NaOH = C,oH,4N,04 + H,0 + CJlT(OH)4COONa. 

Quinine quinate Ouinine Sodium quinate 

Only 100 grams of the original 120 grams of ether-chloroform 
mixture ( = 12 grams of dnchona powder) are in the filtrate. 
This solution contains the alkaloids in 10 grams of bark. 
These 100 grams are extracted with 25 cc. of o.i n-hydrochloric 
add, the alkaloids passing into aqueous solution as hydro- 
chlorides, and the volume is brought to 100 cc. Finally, 
excess of o.i n-hydrochloric acid in 50 cc. (= alkaloids in 5 
grams of bark) of this hydrochloric add solution is determined 
by titration. In these determinations with very dilute hydro- 



QUANTITATIVE ESTIMATION OF ALKALOIDS 



259 



I 

[ 



,oric acid, cinchona alkaloids behave as monacid bases,' 
le forming CsoHa^NjOi.HCl and cinchonine CisHjjNiO.- 

:ci. 

The mean of the equivalent weight of quinine (324) and 
cinchonine (294), that is to say, (324 + 294) divided by 2 =309, 
may be taken as the equivalent weight. This value agrees 
approximately with the actual quantities of these alkaloids 
in cinchona bark. Consequently 1000 cc. of o.i n-hydrochioric 
acid are equivalent to 30.9 grams of cinchona alkaloids. 

Example. — Titration ot 50 cc. of the hydrochloric arid solution of alkaloids, in 

preparing which 12.3 cc. of 0,1 n -hydrochloric add were used, required 3.6 cc. 

: D.I Q-potasEiun hydroxide solution, equivalent to the volume of o.t 

■hydrochloric acid in excess, u.s — 3.6 = g.g cc. of a.i n- hydrochloric acid 

have combined with the allialoids in 5 grams at dnchona bark. The proportion 



Ceo, 



i-HCl:Grams of Alkaloids 



(i - 0.30591) 
0.30591 gram of alkaloids. Consequently 



I grams of alkaloids. 



1 alkaloids at once, 
e to direct sunlight. 



I 



^^lows that 5 grams of barl 

100 grams of bark contain 

Titrate the filtered ether-chloroform solution of c 

The solution should not be exposed for any length of t 
•Otherwise chloroform may give free hydrochloric add 

CHCli + O = COCl, + HCl 

iniiich will neutralLse alkaloids. The decomposition of 0.05 gram of chloro- 
form would give enough hydrochloric add to neutralize o.sj gram of dnchona 
alkaloids. Panchaud* has shown that such chloroform solutions of cinchona 
alkaloids after standing ii hours yield only 80 per cent, of the total quantity of 
alkaloids originally present. 

Hematoxylin, CkHkOi.jHiO, occurs in logn-ood, the heart-wood ot Hema- 
toxyloD campechianum. It usually crystallizes in colorless, shining, quadratic 
prisms containing 3 molecules of water, more rarely in rhombic crystals with i 
molecule of water. It dissolves only slightly in cold water but freely in boiling 
W»ter, alcohol or ether. In contact with air hematoxyiia gradually becomes 
reddish. 

In Aqueous and Alcoholic Cinchona EitiactB. — To determine total alkaloids 
in these preparations, dissolve i grams of the given extract in an Erienmeyer 
fluk, using 5 grams of water and 5 grams of absolute ftlcohol. Add 50 grams of 

'Quinine dihydrochloridc, CmHuNjOi.aHCl, is formed by passing gaseous 
hydrogen chloride over quinine and also by dissolving the monohydrochloride, 
CmHhNiOi.HCI, in strong hydrochloric add with gentle heal. An aqueous 
tolulion of the dihydrochloride has an acid reaction. 

■ Schweiser Wocbenschrift fOr Pharmazie 44, 5S0. 



' 



260 DETECTION OF POISONS 

ether and 20 grains of chloroform, and, after vigorous shaking, 10 cc. <^ sodium 
carbonate solution (i : 3). Shake frequently, and let the mixture stand an hour. 
Then pass 50 grams of the ether-chloroform solution through a dry filter, kept 
well covered. Receive the filtrate in a flask, and distil half the solvent. Pour 
the remainder into a separating funnel, wash the flask three times with 5 cc. 
portions of a mixture of 3 parts of ether and i part of chloroform, and then shake 
the total ether-chloroform solution with 10 cc. of o.i n-hydrochloric add. When 
the contents of the separating funnel are perfectly clear, add enough ether to bring 
the ether-chloroform solution to the surface. Pass the acid solution through a 
small filter moistened with water, and receive the filtrate in a 100 cc. flask. Make 
three more extractions of the ether-chloroform solution with 10 cc. portions of 
water, and pass these extracts through the same filter. Wash the filter with water 
and bring the volume of the filtrate to 100 cc. Finally, measure 50 cc. of this 
solution with a pipette, and add freshly prepared hematoxylin solution, made 
by dissolving a small particle of this substance in i cc. of alcohol. Shake and add 
enough o.i n-potassium hydroxide solution to give the mixture a yellowish color, 
which quickly changes upon vigorous agitation to bluish violet. 

Notes and Calculation. — The alkaloids in the two cinchona 
extracts are set free from their salts by sodium carbonate: 

CwH24N,02 

2 I -h Na,COi = 2CwH,4N,0, -f 2C6H7(OH)4COONa -f H,0 -f CO,. 

CeH7(OH)4COOH 

Quinine Ouinine Sodium 

quinate quinate 

The alkaloids from 2 grams of extract are dissolved in 75 
grams of alcohol-ether-chloroform mixture. Two-thirds of this 
solution, or 50 grams. (= alkaloids in 1.33 grams of extract) 
are used in the determination. The free alkaloids in this por- 
tion pass into aqueous solution as hydrochloride, C2oH^4N202. 
HCl, upon extraction with 10 cc. of 0.1 n-hydrochloric 
acid. The excess of hydrochloric acid in half of this solution 
diluted to 100 cc, that is to say, in 50 cc. (= alkaloids in 0.666 
gram of extract), is finally determined by titration. If 3.7 cc. 
of 0.1 n-potassium hydroxide solution are required, 5 — 3.7 = 
1.3 cc. of 0.1 n-hydrochloric acid have combined with the 
alkaloids in 0.666 gram of cinchona extract. The mean 
equivalent weight of quinine and cinchonine (=309), used in 

the proportion : 

1000 130.9 = 1.3 :x (x = 0.04017) 

shows that 1.3 cc. of 0.1 n-hydrochloric acid correspond to 
0.04017 gram of alkaloids, or 6.03 per cent. The German 



QUANTITATIVE ESTIMATION OF ALKALOIDS 261 

Pharmacopoeia demands this quantity as a minimum for the 
aqueous extract of cinchona bark. 

In the determination of alkaloids in alcoholic cinchona ex- 
tract, titration of excess of o.i n-hydrochloric acid in 50 cc. 
should not require more than 2.3 cc. of o.i n-potassium hy- 
droxide solution. Then 5 — 2.3 = 2.7 cc. of o.i n-hydrochloric 
acid represent the alkaloids in this solution. This extract at 
the minimum must contain 12.55 per cent, of alkaloids. 

Sulphate Method of Estimating Quinine in Mixtures of Cinchona Alkaloids 

(J. Carles)^ 

This method is especially recommended for practical purposes 
because of its accuracy and simplicity. Differences in the 
solubilities of the sulphates of cinchona alkaloids in ammonium 
sulphate solutions form the basis of the method. E. Schmidt 
has found that these sulphates have the following solubilities 
in water at 15°: 

Quinine sulphate i : 800 Cinchonine sulphate i : 65 
Quinidine sulphate i : 100 Cinchonidine sulphate 1 197. 

Guareschi has found quinine sulphate practically insoluble 
in an ammonium sulphate solution, a result which Hille^ has 
confirmed. An addition of 0.0078 gram to the quantity of 
quinine sulphate obtained is necessary on account of the 
quinine sulphate in the 20 cc. of wash water used. 

I. Cinchona Bark. — Place 1 2 grams of finely powdered cinchona bark dried at 
IOC** in an Erlenmcyer flask and add 90 grams of ether and 30 grams of chloro- 
form. Add 10 cc. of sodium hydroxide solution. Shake vigorously at frequent 
intervals for 3 hours. Then add 10 cc. of water, or enough to cause the powdered 
cinchona to gather into baUs after thorough shaking and leave the supernatant 
ether-chloroform layer perfectly clear. After i hour pass 100 grams of the 
ether-chloroform solution through a dry filter kept w^ell covered. Collect the 
filtrate in a dry weighed flask, distil the ether-chloroform and dry the flask at 1 10° 
to constant weight. The increase in the weight of the flask corresponds to the 
total alkaloids in 10 grams of bark. 

Warm the alkaloidal residue in the flask with water and dilute sulphuric acid 

^ Zeitschrift fUr analytische Chemie 9, 467 (1870). 

>W. HiUe (Archiv der Pharmazie 241, 54 (1903)) has reviewed critically the 
various methods that have appeared thus far for the estimation of quinine in pres- 
ence of other cinchona alkaloids. 



262 DETECTION OF POISONS 

and filter the solution. Wash the flask 3 times with water containing sulphuric 
add and pour the wash water through the same filter. Dilute the filtrate to about 
50 CO., heat to boiling and exactly neutralize with ammonia. Cool and after 6 
hours collect upon a weighed filter, or better in a Gooch crucible, the flocculent 
precipitate of quinine sulphate, wash with 20 cc of cold water, dry at zxo^ and 
weigh. 

Add 0.0078 gram to the weight of quinine sulphate found and calculate the 
quantity of quinine in 10 grams of dnchona bark as follows: 

CsoHs4NtOs.HsS04 : CsoHtiNtOt » Quinine sulphate + 0.0078 : x. 

(746) (648) found 

a. Cinchona Extract — Dissolve 3 grams of aqueous cinchona extract in 5 grams 
each of water and absolute alcohol and place in a measuring cylinder. Add 50 cc. 
of ether, 10 cc. of chloroform and, after shaking vigorously 10 cc. of sodium car- 
bonate solution (1:3). Shake at frequent intervals during 3 hours. When two 
layers have formed, bring the ether layer to the 75 cc. mark with more ether. 
Rotate the container carefully and evaporate 50 cc. of the dear ether-chloroform 
layer in a dry weighed flask. Dry i hour at 105° and weigh when cold. The in- 
crease in weight corresponds to the total alkaloids in 2 grams of cinchona extract. 
There should be at least 0.12 gram, or 6 per cent, of alkaloid. 

To determine quinine, pour very dilute sulphuric add over the weighed alka- 
loidal residue in the flask, warm and filter. Rinse the flask several times with 
very dilute sulphuric add, bring the volume to about 50 cc. with water and pro- 
ceed as directed above under cinchona bark. Collect the quinine sulphate in 2 
hours upon a weighed filter, or Gooch crudble. The calculation is the same as 
for dnchona bark. 

Estimatioii of Colchidn in Colchicum Seed and Corms 

(J. Katz and G. Bredemann *) 

Exhaust colchicum seed or conns with 60 per cent, alcohol 
and evaporate 50 grams of this extract to 20 cc. Add 0.5 
gram of solid parafl^e and 20 cc. of water. Warm imtil the 
paraffine is melted and the alcohol has been completely expelled. 
Cool the liquid evaporated to lo-is cc. and pass through a 
moist filter. Melt the paraffine cake upon the water-bath with 
10 cc. of 10 per cent, acetic acid and pour the cold liquid through 
the same filter. Wash the latter, the paraffine cake and the 
dish with water. Saturate the total filtrate with sodium 
chloride and extract first with 20 cc. of chloroform and then with 
10 cc. portions until a few drops of the aqueous liquid show 
scarcely any turbidity with 0.05 n-iodine solution. Pass the 
chloroform solution through a filter moistened with this solvent 

* Pharmazeutsche Zentral-Halle 42, 289 and Apotheker-21eitimg 18, 817. 



QUANTITATIVE ESTIMATION OF ALKALOIDS 263 

and evaporate. To expel chloroform retained by the colchicin, 
dissolve the residue in a little water and j&lter. Evaporate the 
solution in a weighed dish and dry the residue over sulphuric 
acid to constant weight. 

Note. — Using this method, Bredemann obtained the following quantities of 
colchicin: 

In seed 0.46 -0.13 per cent. 

In corms 0.032-0.06 per cent. 

In fresh flowers 0.6 per cent. 

In dry flowers 1.8 per cent. 

Alkaloids in Pomegranate Bark 

Pomegranate bark, the bark of Punica Granatum, contains 
the following four alkaloids: 

Pelletierine, CgHuNO, Methyl-pelletierine, C9H17NO, 

Isopelletierine, CsHuNO, Pseudo-pelletierine, CgHisNO. 

According to Piccini there is still another alkaloid in the bark of 
pomegranate root isomeric with methyl-pelletierine and there- 
fore called isomethyl-pelletierine. 

Ciamician and Silber have determined the structure of 
pseudo-pelletierine which they call n-methyl-granatonine. 
Pseudo-pelletierine (I) is a ketone which, upon treatment with 
sodium amalgam or with sodium and alcohol, adds two atoms 
of hydrogen and passes into the corresponding secondary 
alcohol, n-methyl-granatoline (II). Chromic and sulphuric 
acids oxidize the latter to n-methyl-granatic acid (III). Nitro- 
gen can be eliminated by exhaustive methylation and the final 
product is normal suberic acid (IV) : 

I. H,C— CH— CH, II. H,C— CH — CH, 



H,C N.CH,CO H,C N.CH,CH.OH 

T I I +2H — > I I I - 

H2C— CH — CH, H,C— CH — CH, 

Pseudo-pelletierine ■- n-methyl-granatolin 

n-roethyl-granatonine 

ra. H,C— CH— COOH IV. H,C— CH,.COOH 

I I I 

H,C N.CH, — ♦ H,C 



H,C— CH— CH,.COOH H,C— CH,.CH,.COOH 

n-methyl-granatic acid Normal suberic acid 



[,C— ( 



264 DETECTION OF POISONS 

Estimation of Alkaloid in Pomegranate Bark 
(German Pharmacopoeia) 

To determine total alkaloids, pour 90 grams of ether and 30 grams of chloro- 
form upon 12 grams of rather finely groimd pomegranate bark, dried at 100° and 
placed in an Erlenmeyer flask. Shake vigorously and add 10 cc. of a mixture of 

2 parts of sodium hydroxide solution and i part of water. Let the mixture stand 

3 hours, shaking vigorously at frequent intervals. Then add 10 cc. of water, 
which will cause the powder to gather into balls after vigorous shaking, and leave 
the supernatant ether-chloroform solution perfectly clear. After an hour, pass 
ICO grams of the dear ether-chloroform solution through a dry filter, kept well 
covered and receive the filtrate in a separating funnel. Extract this filtrate 
with 50 cc. of o.oi n-hydrochloric add and pass this add solution, when perfectly 
dear, through a small filter moistened with water into a 100 cc. flask. Make 
three more extractions with 10 cc. portions of water, and pass these extracts 
through the same filter. Wash the filter with water and dilute the total solution 
with water to 100 cc. Place 5c cc. of this solution in a 200 cc. flask with 50 cc 
of water and enough ether to make a layer about i cm. thick. Add 5 drops of 
iodeosine solution and enough o.oi n-potassium hydroxide solution, shaking 
vigorously after each addition, to give a pale red color to the lower aqueous 
solution. 

Calculation. — ^The 100 grams of filtered ether-chloroform solution correspond 
to 10 grams of bark. The alkaloids are transferred from this ether-chloroform 
solution to 50 cc. of o.oi n-hydrochloric add which are diluted with water to 
100 cc. The excess of hydrochloric add in 50 cc. of this solution ( = alkaloids 
from 5 grams of bark) is determined by titration. If, for example, 11 cc. of o.oi 
n-potassium hydroxide solution are used, then 25 — 11 = 14 cc. of o.oi n-hydro- 
chloric add have combined with the alkaloids in 50 cc. of the solution. If the 
mean of the equivalent weights of pdletierine (141) and pseudo-pelletierine (153), 
or 147, is used in the calculation, 1000 cc. of o.oi n-hydrochloric add neutralize 
1.47 grams of the mixed alkaloids. According to the proportion 

1000 : 1.47 =» 14 : X (x = 0.02058) 

5 grams of pomegranate bark contain 0.02058 gram of alkaloids which corre- 
sponds to an alkaloid content of 20 X 0.02058, or 0.41 per cent. The German 
Pharmacopoeia demands this quantity of alkaloids in pomegranate bark as a 
minimum. 

Estimation of Caffeine in Coffee, Tea, Cola Nuts and Guarana 

(Literature) 

A. Hilger and A. Juckenack. — Zur Bestimmung des Kaffelns in Kafifee und 
Tee. Forschungsberichte tiber Lebensmittel imd ihre Beziehungen zur Hygiene 
4i 49-50; C^ 1897 I, 775 and also 4, 145-154 and C 1897 II, 233. 

H. Trillich and H. G5ckeL — Bdtrage zur Kenntniss des Kaffees imd der 
Kaffeesurrogate. Forschimgsberichte Qber Lebensmittel und ihre Beziehungen 
zur Hygiene 4, 78-88 and C 1897 I, 1248. 

* C = Chemisches Zentralblatt. 




QUANTITATIVE ESTIMATION OF ALKALOIDS 



265 



L. Graf.^Ueber Zusammcnbang von KaUdngebalt und Qaatiiat bei cMdcs- 
ischen Tec. Forschungsbcrkhte liber Lebensmittel und ihre Beziehungen lur 
Hygieoc 4, 88-89, and C 1897 I, '249- 

A. FoTster and R. Riechelmaim. — Zur Bestimmung des KalTetns Im KaSte. 
Zeitsclirift ftir dffentliche Chcmie 3, 129-131 and C 1897 I, 1)59. 

C. C. Keller.— Die Bcslinimung des Kaffelns im Tee. Berichte der Deutschen 
pharmnceulischen Gcscllsthufl 7, 105-111 (1897) and C i8g7 I, 1134. 

A. Forater and A Rieclieliuaiin. — Zur Bestimmung des KalTdns im KaSee. 
(Enlgegnung.) Zeitschrift fUr Sffectliche Chemie 3, 235-336 and C 1897 II, 
«&■ 

E. Tas^y. — Uebet eiD neues Verfabrenzur Bestimmung des KaSelns im Kaffee. 
Bulletin dc la Sociftfi chimiquc. Paris. (3) 17. 766-768 and C 1897 II, 644- 

K. Dieterich. — Uebec die Wcrthbestimmung der Kolanuss und dea Kolaex- 
traktes. Vortrag auf der Naturforschcrversammlung in Braunschweig gehaltco. 
Pharmaceutischc Zeitung 41, 647-650 and C 1897 11, 977. 

H. Bnmner and H. Leins. — Uebcr die Trentiutig und quaniiiative Bestim- 
mung des Kaffdns und Theobromins. Schweizer Wochenscbrifc fOr Pharmacie 
36, 301-303 and C 1898 ri. $12. 

J. Gadomer. — Uebet Kaffelnbcstimmungen in Tec, Kaffee und Kola. Archiv 
der Pharmade 337, 58-68 and C 1899 I, 713. 

F. Katz. — Ueber die quantitative Bestimmung des Kaffelns. Berichte der 
Deulschen pharmaceutischen GeseUschaft u, 250 (1902). 



Vopoi 






C. C. Keller's Method. — Pour 120 grams of chloroform 
in 6 grams of dry, unbroken tea leaves^ in a wide-mouth 
separating funnel. In a few minutes, add 6 cc. of ammonium 
hydroxide solution (10 per cent. HjN), and at frequent intervals 
shake vigorously during 30 minutes. Then let the separating 
funnel stand at rest, until the solution is perfectly clear and the 
tea leaves have absorbed all the water. This may require 
3-6 hours, or even longer, depending upon the variety of tea. 
Pass 100 grams of clear chloroform extract, representing 5 
grams of tea, through a small filter moistened with chloroform, 

ve the filtrate in a small, weighed flask and distil the chloro- 
upon the water-bath Pour 3-4 cc. of absolute alcohol 

the residue. Heat upon the water-bath to remove alcohol 
and expel alcohol vapor with a hand bellows. In a few minutes 
the caffeine will be dry and at the same time free from im- 

'When a. wide-mouth separating funnel cannot be obtained, triturate the 
tea leaves somewhat, solely to facilitate their removal from the separating funnel 
after extraction. Finely powdered tea is not only unnecessary, but even ob- 
jectionable, because the extracts have a much deeper color, and the yield of 
tfidne is not increased. 



266 DETECTION OF POISONS 

prisoned chloroform. In a measure also, this treatment with 
alcohol separates caffeine from extraneous chlorophyll. The 
latter adheres to the bottom and side of the flask, whereas 
cafifeine forms a white incrustation upon it. Caffeine thus 
obtained is usually impure from small quantities of ethereal 
oil, fat, vegetable-wax and principally chlorophyll. Conse- 
quently, it must be purified. Set the flask upon a boiling water- 
bath and pour a mixture of 7 cc. of water and 3 cc. of alcohol 
over the crude caffeine, which upon being shaken will pass into 
solution almost immediately. Then add 20 cc. of water, 
stopper the flask and shake vigorously. The chlorophyll will 
form lumps and the solution will filter easily. Pass the caffeine 
solution through a small filter moistened with water, wash flask 
and filter with 10 cc. of water, evaporate the total filtrate in 
a weighed glass dish to dryness upon the water-bath and 
weigh the residue of nearly pure caffeine. The weight of this 
residue multiplied by 20 will give the percentage of caffeine in 
the tea. 

Notes. — Ammonia causes tea leaves to swell considerably, 
and at the same time combines with the tannic acid present. 
Caffeine is set free and dissolved by chloroform. The color of 
the chloroform extract depends upon the variety of tea. Black 
teas (Pekoe, Souchong and Congo) give clear, pale green to 
yellowish green solutions. Teas not so black, or green teas, 
give darker and more brownish green solutions. In assaying 
those varieties of tea, which probably contain a small quantity 
of caffeine, take 12 grams and extract with 150 grams of 
chloroform. C. C. Keller has shown that the best and most 
expensive varieties of tea contain most caffeine. The average 
percentage of caffeine, based upon 50 assays of tea, was found 
to be 3.06. A green tea gave the smallest yield, namely, 
1.78 per cent, of caffeine; and a Pekoe tea the highest yield, 
namely, 4.24 per cent, of caffeine. 

J. Gadamer states that Keller's method of estimating caffeine 
in tea is applicable also to coffee and cola preparations. Keller's 
method is especially useful for roasted coffee. The caffeine, 
though somewhat brown, is always sufficiently pure. 



QUANTITATR-E ESTIMATION OF ALKALOIDS 



267 



I 
[ 



I 



2. Hilger-Juckenack Method. — Macerate 20 grams of finely 
ground coffee, or triturated tea, with 900 grams of water for 
several hours in a large beaker at room temperatures. Then 
boil thoroughly and replace the water lost by evaporation. 
Raw coffee requires 3 hours and roasted coffee and tea 1.5 
hours. Cool somewhat (60 to So") 
and add 75 grams of aluminum 
acetate solution (see note, page 
282) and gradually, while stirring, 
1.9 grams of acid sodium carbon- 
ate. Boil about 5 minutes and 
bring the total weight when cold 
to 1020 grams. Filter 750 grams 
(=15 grams of original material) 
and add to the clear filtrate 10 
grams of precipitated and pow- 
dered aluminium hydroxide and 
some filter paper made into a 
magma by agitation with water. 
Stir frequently and evaporate 
upon the water- bath. Thoroughly 
dry the residue in an air-closet at 
100", and extract for 8-10 hours 
with pure tctracblorom ethane 
(CCli), using a Soxhlet apparatus 
(Fig. 33). Tetrachloromethane, 
which is always colorless, is finally 
distilled and the residue of per- 
fectly white caffeine is dried at"^ 
100° and weighed. The results 
thus obtained are usually accepted 
without question. But if an absolutely accurate result is 
required, nitrogen in the crude caffeine may be determined by 
KJeldahl's method. The quantity of anhydrous caffeine is 
calculated on the basis of this analysis. One cc, of o.i n-oxalic 
acid represents 0.00485 gram of anhydrous caffeine. Com- 
mercial tetrachloromethane is usually impure and cannot be 




J. — Soxhlet Apparalua. 



268 DETECTION OF POISONS 

used directly in the extraction. It should be shaken 3-4 times 
with sodium carbonate solution, then several times with water, 
dried over fused calcium chloride and distilled fractionally. 
It boils at 76-77°. 

3. Trillich-Goeckel Modification of Hilger's Metbod. — 
Exhaust 10 grams of finely ground coflfee with water. This 
will require 3 extractions with boiling water, using 200 cc. 
portions and heating each for 30 minutes. Combine thejUtered 
extracts, cool and dilute to 495 cc. Add 5 cc. of basic lead ace- 
tate solution, shake thoroughly, filter 400 cc. and pass hydrogen 
sulphide through the filtrate. Dilute this filtrate to 500 cc, 
shake thoroughly and again filter 400 cc. This filtrate will 
represent 6.4 grams of coffee. Concentrate this filtrate 
(400 cc.) upon the water-bath, and, after addition of i gram of 
magnesium oxide and sand, evaporate to dryness. Triturate 
the residue and extract for 30 hours with acetic ether, using a 
Soxhlet apparatus. Evaporate the acetic ether extract in a 
Kjeldahl flask, or distil the solvent, and determine nitrogen in 
the residue by Kjeldahl's method. One gram of nitrogen repre- 
sents 3.4643 grams of caffeine. Crude caffeine is easily de- 
composed by the acid used in the Kjeldahl process and by mer- 
curic oxide. Roasted coffee may easily give too much caffeine 
by this method, because the bases formed by roasting coffee, 
pyridine for example, are also extracted by acetic ether. 

4. Triliich-Goeckel Modification of Socolof s Method. — 
Put 10 grams of finely ground, dried coffee into a separating 
funnel, provided with a plug of glass wool for a filter, and 
moisten with ammonium hydroxide solution. Let the mixture 
stand for 30 minutes, extract for 12 hours with 200 cc. of acetic 
ether and shake frequently. Filter, and wash three times with 
50 cc. portions of acetic ether. Distil the acetic ether upon the 
water-bath and boil the residue with milk of magnesia. Filter, 
and evaporate the filtrate to dryness upon the water-bath. 
Dissolve the residual caffeine in acetic ether or chloroform. 
Filter this solution into a weighed dish or Kjeldahl flask. 
Evaporate the solvent and weigh the caffeine, or calculate it 
from the percentage of nitrogen. The latter method is the more 



» 



I QUANTITATIVE ESTIMATION OF ALXALOIDS 269 

accurate. According to C. Wolffs the residue from the acetic 
ether or chloroEorm extract should not be accepted as pure 
caffeine. Determination of nitrogen in this residue by Kjeldahl's 
method is the most reliable way of estimating caffeine in the 
extract, 

5. E. Katz's Method of Estimating Caffeine. — This method 
is based upon the fact that chloroform will extract caffeine 
quantitatively from a solution which is ammoniacal, or faintly 
add with hydrochloric add. 

Shake lo grams of powdered coffee, or tea, for 30 minutes 
with 200 grams of chloroform and 5 grams of ammonium 
hydroxide solution. When the liquid has settled, filter 150 
grams of the chloroform solution through a Sander's filter 
which will give a perfectly bright filtrate free from water. Dis- 
til the chloroform completely and dissolve the residue with 
gentle heat in about 6 cc. of ether. Add so cc. of 0.5 per cent, 
hydrochloric add and, in an assay of coffee, also 0.2-0.5 gram 
of solid paraftine. Evaporate the ether and filter the cold, 
aqueous solution. Wash the flask and filter paper a few times 
with small portions of 0.5 per cent, hydrochloric add. Finally 
extract the total aqueous hydrochloric add solution four times 
with 20 cc. portions of chloroform. Distil the filtered chloro- 
iorm extract, dry the residue and weigh. This residue will 
consist of nearly pure caffeine. J. Katz found the following 
percentages of caffeine: 

Caffeine Average 

Raw Coffee Beans o.g -1.37 per cent. t.14 per cent. 

Dried Cola Nuts 1.51-1,54 per cent, 1.68 per cent. 

Bkck Tea 1.51-3-56 percent. 3.07 per cent. 

Guarana 2.83-4,74 per cent. 4,08 per cent. 

J. Katz recommends the following method for estimating 
caffeine in matfi or Paraguay tea: 

Treat finely triturated tea with ammonium hydroxide solu- 
tion and chloroform, as described above, and dissolve the chloro- 
form residue in ether. Add water to the ether and evaporate. 
Warm the aqueous solution 10 minutes upon the water-bath 

1 Zeitschrifl far iJEfenllkhe Chemie 11. 186. 



I 
I 



270 DETECTION OF POISONS 

with 2 cc. of lead hydroxide suspended in water (i :2o). If it 
is very difficult to get a clear filtrate from this liquid, add a 
little calcined magnesium oxide. This treatment usually gives 
a filtrate, which is perfectly clear when cold, and but slightly 
colored. Chloroform extracts quite pure caffeine itbia this 
solution. By this method mat6 yields 0.3-1.6 per cent- of 
caffeine, the average being 0.71 per cent. 

6. K. Dieterich's Method of Estimating Total Alkaloids 
(Caffeine and Theobromine) in Cola Nuts. — Moisten 10 grams 
of finely grated cola nuts with a little water, mix with 10 grams 
of granulated, unslaked lime and extract with chloroform in a 
Soxhlet apparatus for 45 minutes. Evaporate the extract 
almost to dryness and dissolve the residue with gentle heat 
in 20 cc. of normal hydrochloric add. Filter and dilute to 100 
cc. Add ammonium hydroxide solution in large excess to this 
filtrate, shake at frequent intervals during 15 minutes and 
extract three times with 20 cc. portions of chloroform. Evapo- 
rate this chloroform solution in a weighed flask and dry the 
residut\ which usually consists of perfectly pure caffeine, at 
100^ to constant weight. 

This method may also be used in estimating caffeine in 
Paraguay tea. Mix the finely ground material with unslaked 
lime, and extract with chloroform, in a Soxhlet apparatus. Tea 
gives pure, white caffeine free from chlorophyll. 

Estimation of Alkaloids in IpecMc 
IjHKac has been shown* to contain three alkaloids: 

Ccphaeline, C1SH40N1O4, Emetine, CMH44N1O4, 
Psvcho trine. 

The composition of the last alkaloid is unknown. This drug 
acts as an expectorant and emetic, because of cephjdine and 
emotino. Psvchotrine is said not to possess these properties. 
Thorctore, in assaxnng ipecac for medicinal purposes, only the 
l>orccntagc of the first two alkaloids need be estimated. The 
equivalent weights of these two alkalcads (cephjdine 254 and 

^ rrerioh$ atkI dc risentJis TipBS« Aixjiiv der Fkazmftcie^ 1902, Heft 5 a»l 6. 



QDANTITATIVE ESTIMATION OF ALKALOIDS 
; 248) £ 



271 



I 



emetine 248) are so nearly the same, that the mean of the two 
(241) may be used as the factor. 

Procedure. — Put 6 grams of finely powdered root in a dry 
Erlenmcyer llask and shake with 60 grams of ether. Then add 
S cc. of ammonium hydroxide solution, or 5 cc. of sodium 
carbonate solution (1:3), and shake frequently during an hour. 
Add 10 cc. of water and, after shaking vigorously, filter 50 
grams of the ether extract into a small flask. Evaporate half 
the ether upon the water-bath, and extract the remainder in a 
separating funnel with 10 cc. of o.i n-hydrochloric acid. Pass 
the acid solution through a small filter into a 200 cc. flask. 
Make two more extractions of the ether with 10 cc. portions of 
water, and pass these through the same filter. Bring the 
volume of the acid solution to 100 cc, and then add enough 
ether to form a layer about i cm, thick after thorough agitation. 
Add 5 drops of iodeosine solution (1:250), and titrate excess of 
hydrochloric acid with o.i n-potassium hydroxide solution. 
The number of cc- of 0.1 n-hydrochloric acid, combined with 
the alkaloids, multiplied by 0.0241 gives the quantity of eme- 
tine and cephffiline in s grams of ipecac. 

To estimate these alkaloids gravimetrically, shake vigorously 
the ether solution of the alkaloids (50 grams = 5 grams of root) 
in a separating funnel with 5 cc. of dilute hydrochloric acid 
and 10 cc. of water. Transfer the acid solution to another 
separating funnel. Make two more extractions of the ether 
with 10 cc. portions of water and add these to the acid extract. 
Add s cc. of ammonium hydroxide solution to the acid extract 
and shake vigorously with sograms of ether. Remove the aque- 
ous layer and filter 40 grams of the ether solution into a weighed 
flask. Evaporate the ether and weigh the flask after drying 
for an hour at 100°. This will give the quantity of emetine and 
cephseline in 4 grams of root. 

Test for Cephseline. — This reaction is very characteristic of 
this alkaloid. Froehde's reagent dissolves pure cephjeline, as 
the free base, almost without color. A trace of hydrochloric 
acid, or better sodium chloride, added to this solution produces 
an intense blue color. Pure emetine gives no color with 



272 DETECTION OF POISONS 

Froehde's reagent, nor when sodium chloride is added. This 
test for cephaeline may be made with the ether residue. 

The method of estimating alkaloids in ipecac, prescribed by the German 
Pharmacopoeia, is the same as that for cinchona bark. Use 12 grams of finely 
powdered root dried at 100°, but in ascertaining excess of add use iodeosine, 
and not hsmatoxylin, as the indicator. Finally, measure with a pipette 50 
cc. of the proper solution having a volume of 100 cc, place in a 300 cc. flask 
and add about 50 cc. of water and enough ether to make a layer z cm. thick. 
Add 5 drops of iodeosine solution and enough o.oi n-potassiimi hydroxide 
solution, shaking thoroughly after each addition, to give the lower aqueous 
layer a pale red color. This should require not more than 20 cc of alkaline 
solution. 

Estimation of Nicotine in Tobacco 

1. R. Kissling's^ Method. — First remove the ribs and then 
cut the tobacco leaves into small pieces. Dry 1-2 hours 
('50-60°), and then reduce to a uniform, coarse powder. 
Triturate 20 grams of this powder with 10 cc. of dilute, alcoholic 
sodium hydroxide solution (6 grams of sodiimi hydroxide 
dissolved in 40 cc. of water and 60 cc. of 95 per cent, alcohol). 
Transfer this moist powder to a paper thimble and extract 
2-3 hours with ether in a Soxhlet apparatus. Carefully distil 
the ether solution so that a portion of the solvent remains. Add 
50 cc. of very dilute sodium hydroxide solution (4 grams of 
sodium hydroxide in 1000 cc. of water) to the residue and distil 
with steam. Begin introducing steam after the nicotine solu- 
tion has been boiling several minutes. Collect about 400 cc. of 
distillate, that is to say, continue distillmg until the distillate is 
no longer alkaline. Mix well, add a few drops of rosolic add 
solution to the distillate, and titrate nicotine with o.i n-sulphuric 
or oxalic acid until the red color has just disappeared. 

Calculation. — Although nicotine, C10H14N2 (162), as a di-acid 
base can combine with two equivalents of acid, it behaves upon 
titration, with rosolic acid or iodeosine as indicator, as if it 
were a monacid base with the equivalent weight 162. 1000 cc. 
of o.i n-acid consequently correspond to 16.2 grams of nicotine. 

2. C. C. KeUer's^ Method. — Pour 60 grams of ether and 60 

*2^itschrift fiir analytische Chemie 34, 1731 and 21, 76. 

* Berichte der Deutschen pharmazeutischen Gesellschaft 8, 145 (1898). 



QUANTtTATrV-E ESTIMATION OF ALKALOIDS 



273 



i 



grams of petroleum ether over 6 grams of dry tobacco in a. zoo 
CO. Erlenmeyer flask. Add lo cc. of 20 per cent, aqueous 
potassium hydroxide solution and let the mixture stand half an 
hour, shaking vigorously at frequent intervals. After the Uquid 
has stood at rest 3-4 hours, pour 100 grams of ether solution 
through a small, plaited filter and receive the filtrate in a 200 cc. 
Erlenmeyer flask. Nicotine is in solution together with a little 
ammonia, which must be removed before titration. By means 
of a hand bellows and a glass tube reaching to the bottom of the 
flask force a current of air through the solution, so that there is 
considerable agitation. It requires about a minute and a half 
to expel all ammonia. At the same time 8-10 grams of ether 
evaporate. Add 10 cc. of alcohol, a drop of i per cent, iodeo- 
sine solution and 10 cc. of water to the ammonia-free solution. 
Stopper the flask and shake vigorously. Nicotine and iodeo- 
sine dissolve in the water which has a red color. Adda slight 
excess of o.i n-hydrochloric acid, enough to discharge the color, 
and titrate excess of acid with o.i n-potassium hydroxide solu- 
tion. The quantity of nicotine in tobacco shows a wide varia- 
tion and ranges from 0.6 to 4.8 per cent. 

3. J. Toth's' Method.— According to Toth two sources of 
error in C. C, Keller's method lead to low results. An aqueous 
potassium hydroxide solution retains variable quantities of 
nicotine and a current of air passed through an ether solution 
of nicotine volatilizes some of this alkaloid. Therefore Toth 
recommends the following procedure: 

Mix 6 grams of air-dried tobacco with 10 cc. of 20 per cent, 
sodium hydroxide solution in a porcelain dish. Add gypsum 
until the mixture is like powder. Extract thoroughly with 
100 cc. of ether-petroleum ether mixture (i ; i) and after i hour 
pipette off as quickly as possible 25 cc. of the solvent. Add 
40-50 cc. of water, i drop of lodeosine solution and an excess of 
O.I n-sulphuric acid. Determine excess of acid by titration 
with O.I n-sodium hydroxide solution. The ether-petroleum 
mixture takes up at most 0.0005 gram of ammonia. 



' Chemisches Zenlralblatl, i 



274 DETECTION OF POISONS 

Estimatioii of Hydrastine in Fluid Extraet of Hydrastis 

(German Pharmacopceia) 

Evaporate 15 grains of fluid extract of hydrastis to about 5 grams in a weighed 
dish upon the water-bath, and wash the residue into an Erlenmeyer flask with 
about 10 cc. of water. Add 10 grams of petroleum ether, 50 grams of ether and 5 
grams of ammonium hydroxide solution. Let the mixture stand an hour, shaking 
vigorously at frequent intervals. Then pass 50 grams of the dear ether solution 
through a dry filter into a separating funnel. Add 10 cc. of a nuxture, composed 
of I part of hydrochloric add and 4 parts of water, and shake the solution 
vigorously several minutes. When the liquids have separated dear, run the 
add solution into an Erlenmeyer flask. Make two more extractions of the ether 
with 5 cc. portions of water containing a few drops of hydrochloric add, and add 
these to the first extract. Add to the total extract excess of ammonium hydroxide 
solution and 50 grams of ether. Let the mixture stand an hour, shaking vigor- 
ously at frequent intervals. Pass 40 grams of the clear ether solution through 
a dry filter and collect the filtrate in a weighed, dry flask. Distil the ether, 
dry the residue at 100° and weigh when cold. The residue should weigh at 
least 0.2 gram. 

Notes. — Additional Information about hydrastine is given 
on page 112. Ammonia, added to an aqueous solution of the 
residue from hydrastis extract (15 grams), sets the alkaloids, 
hydrastine and berberlne, free from their salts. The ether- 
petroleum benzine mixture dissolves hydrastine but not ber- 
berlne, the latter being nearly insoluble In this mixed solvent. 
But phytosterln, which Is always present in hydrastis extract, is 
dissolved. Only 50 grams (= hydrastine in 12.5 grams of 
extract) of the original 60 grams of ether-petroleum benzine 
mixture are used. Hydrastine is extracted from the solvent by 
agitation with dilute hydrochloric acid and dissolved in the 
acid solution as hydrochloride. The alkaloid is then precipi- 
tated from the acid solution by ammonia and dissolved in 50 
grams of ether: 

C2iH2iNO«.HCl + (H4N)0H = C«H8,N0« + H,0 -h (H4N)C1. 

Hydrastine Hydrastine 

hydrochloride 

The hydrastine in 40 grams of the ether solution (=10 grams 
of original extract) Is finally weighed. Good extract of hydras- 
tis should contain 2-2.5 P^^ cent, of hydrastine. 

When the ether-petroleum benzine solution of hydrastine and 
phytosterln Is extracted with dilute hydrochloric acid, the 
alkaloid passes into the acid solution free from phytosterin. 



QUANTITATIVE ESTIMATION OF ALKALOIDS 



275 



crolonate Method of Estumttuig Hydiasdne i 
Extract 



Hydrastis Root and 



(H. Matches and O. Raromstedt)' 



matiDn of Berberine.— ^This alkaloid has only a slight phyaiologicaJ action. 
To determine approximately the quantity present in hydrastis extract, add >□ 
grams o[ dilute sulphuric acid (1:5) to 10 grama of the extract and let the miicture 
stand for 24 hours at as low a temperature as possible. Crystallization of ber- 
berine as the dilQcultly soluble acid sulphate. CitHuNOf.HiSOi, is almost com- 
plete. Filter in a Gooch crucible with suction, washing lirst with a little water 
containing sulphuric acid and then with pure water. Dry at 100° to constant 
weight. (E. Schmidt.) 

W. Meine' has found that the crystalline deposit, frequently seen in hydraatis 
extract, consists mostly of berberine mixed with a little phytosterin. This 
deposit is said to conta.in only tiaccs of hydrastine. 

^picrol 

r The German Phannacopceia requires the estimation of 
hydrastine but not of the physiologically inert substances, 
berberine and phytosterin, also present in Hydrastis prepara- 
tions. Ether-petroleum benzine mixture, used as a solvent, 
dissolves phytosterin and hydrastine but not berberine. Di- 
lute hydrochloric acid extracts hydrastine but leaves phytos- 
terin in the ether mixture. 

Estimation of hydrastine by means of picrolonic acid appears 
simpler than by the method of the Pharmacopceia, because 
picrolonic acid does not precipitate phytosterin and therefore 
hydrastine is not mixed with this impurity, Matthes and 
Rammstedt obtained nearly pure hydrastine picrolonate from 
hydrastis extract, melting at 220-325°. The picrolonate pre- 
pared from pure hydrastine, CjiHiiNOB.CioHgN406, melts at 

225°. 

I. In Fluid Extract of Hydrastis. — Evaporate 15 grams of 
fluid extract to about 5 grams in an Erlenmeyer flask upon the 
water-bath. Add 10 cc. of water; 10 grams of petroleum 
benzine, 50 grams of ether and 5 grams of ammonium hydroxide 
(10 per cent. NHj). Shake vigorously for lo minutes. After 
"le mixture has stood for 20 minutes, pour 40 grams of the ether- 

' Zeilschrift des allgemcinen osterreichischen Apothefcer-Vereins 55. 494. 

■ Further information about picrolonic add and itsuse in predpitating alkaloids 

given on page 346. 



276 DETECTION OF POISONS 

benzine extract through a double, creased filter and evaporate 
about one-half in a beaker. Then add lo cc. of o.i n-picrolonic 
add solution. After 24 hours collect the hydrastine picrolonate 
in a weighed Gooch crucible, wash with 2 cc. of an alcohol-ether 
mixture (1:3), dry for 30 minutes at 105® and weigh. 

2. In Hydrastis Root — Shake 6 grams of powdered root 
vigorously for 30 minutes with 50 grams of ether, 10 grams of 
petroleum ether and 6 grams of ammonium hydroxide (10 per 
cent. NHs) . Then add 6 grams of water and shake until the 
upper layer of liquid is clear. Quickly filter 50 grams of the 
ether-petroleum benzine extract and evaporate about one-half 
in a beaker. Then add 5 cc. of o.i n-picrolonic acid. After 24 
hours filter the picrolonate precipitate and wash with i cc. of 
alcohol-ether mixture (1:3). Otherwise the procedure is the 
same as described in i . 

In the calculation use the formula of hydrastine picrolonate 
(Mol. Wt. 647) given above. 

Estimation of Morphine in Opium and Pharmaceutical Preparations 

(German Pharmacopoeia) 

In Opium. — Triturate 6 grams of rather finely powdered opium with 6 grams of 
water. Wash the mixture into a weighed, dry flask with water and add enough 
more of this solvent to bring the weight to 54 grams. Shake frequently and let 
the mixture stand an hour. Pour upon a piece of dry linen and express the 
liquid. Pass 42 grams of this extract through a dry, plaited filter (10 cm. in di- 
ameter) into a dry flask. Add 2 grams of sodium salicylate sqlution (i : 2) to this 
filtrate and shake vigorously. Filter 36 grams of the clear solution through a dry, 
plaited filter (10 cm. in diameter) into a small flask. Mix this filtrate by gentle 
agitation with 10 grams of ether, and add also 5 grams of a mixture consisting of 
1 7 grams of ammonium hydroxide solution and 83 grams of water.* Stopper the 
flask, shake vigorously for 10 minutes and let the mixture stand at rest 24 hours. 
Then decant the ether layer as completely as possible upon a smooth filter 
(8 cm. in diameter). Add 10 grams more of ether to the residual, aqueous liquid 
in the flask, shake gently for a few minutes and again pour the ether layer upon 
the filter. Then after all the ether solution has passed through, pour the 
aqueous solution upon the filter, and disregard crystals adhering to the side of 
the flask. Wash filter and flask three times with 5 cc. portions of water satu- 
rated with ether. When the filter has drained thoroughly, dry the morphine 
crystals and dissolve in 25 cc. of 0.1 n- hydrochloric acid. Pour this solution 
into a 100 cc. flask, carefully wash filter and flask with water and finally dilute 
the solution to 100 cc. Measure 50 cc. of this solution into a 200 cc. flask, add 



QUANTITATIVE ESTIMATION OF ALKALOIDS 277 

50 cc. of water and enough ether to form a layer z cm. thick. Add 5 drops of 
iodeosine solution and enough o.i n-potassium hydroxide solution, shaking vigo- 
rously after each addition, to produce a pale red color in the lower aqueous layer. ^ 

Notes and Calculation. — Most of the opium alkaloids are 
combined with meconic (see page 205) and sulphuric acids. 
Ammonium hydroxide, added to an aqueous opium extract, 
sets the alkaloids free from their salts: 

(C17H1.NO,), OH 

C»HO,(OH)(COOH), + 2(H4N)OH = 2Ci7Hi«NO, + 2H,0 + C»HO, 



I 



. :00NH4)f 

Morphine meconate . Morphine Ammonium 

meconate 

The ether used dissolves all opium alkaloids except morphine 
which having once become crystalline is insoluble in this 
solvent. Saturated sodium salicylate solution precipitates 
resinous and greasy substances from the filtered aqueous opiimi 
extract and also narcotine which next to morphine is present in 
opium in largest quantity. 

The moprhine from 6 grams of opium is in 54 grams of filtered 
aqueous extract. After the second filtration only 36 grams of 
this extract are used (= morphine from 4 grams of opium). 
Morphine, precipitated by ammonium hydroxide from these 36 
grams of extract, is dissolved in 25 cc. of o.i n-hydrochloric acid 
as hydrochloride, C17H19NO3.HCI. This solution is then diluted 
to 100 cc. and excess of acid in 50 cc. of this hydrochloric acid 
solution (= morphine from 2 grams of opium) is determined. 

Morpine being a monacid base has the same molecular and 
equivalent weights = C17H19NO3 = 285. Therefore 1000 cc. 
of O.I n-hydrocMoric acid = 28.5 grams of morphine. 

Example. — Titration with o.i n-potassium hydroxide solution has shown that 
there are 4.1 cc. of 0.1 n-hydrochloric acid in 50 cc. of the hydrochloric acid 
solution of morphine. There remain therefore 12.5 — 4.1 = 8.4 cc. of the 0.1 
n-add originally present now combined with the morphine from 2 grams of opium. 
According to the proportion 

Cc. O.I n-HCl : Grams morphine 

1000 : 28 .5 = 8.4 : X (x = 0.2394) 

^ The German Pharmacopoeia demands that not more than 5.4 cc. nor less than 
4.1 cc. of O.I n-alkaline hydroxide solution, corresponding to a morphine-content 
of 10-12 per cent, shall be used to produce this color. 



278 DETECTION OF POISONS 

8.4 cc. of O.I n-acid correspond to 0.2394 gram of morphine. Consequently the 
opium contains 50 X 0.2394 — 11.97 per cent, of morphine. This is the maxi- 
mum quantity of morphine allowed in opium by the German Pharmacopceia. 

a. In Extract of Opium* " Dissolve 3 grams of opium extract in 40 grams of 
water, add 2 grams of sodium salicylate solution (1:2), shake vigorously, pass 30 
grams of clear solution through a dry filter (10 cm. in diameter) and coUect 
in a dry flask. Mix this filtrate with 10 grams of ether by rotating the flask and 
add also 5 grams of a mixture of 1 7 grams of anmionium hydroxide and 83 grams 
of water.'' Continue the assay as directed above in i (Opium) from the point 
marked with an asterisk. 

Calculation. — Only 2 of the 3 grams of opium extract wdghed are used in the 
determination, since only 30 grams of the original 45 grams of solution (3 grams 
of extract + 2 grams of sodium salicylate solution + 40 grams of water) are 
filtered. The morphine obtained from these 2 grams of extract is dissolved in 25 
cc. of o. I n-hydrochloric acid and the volume is then brought to 100 cc. The titra- 
tion uses 50 cc. of this solution which contains the morphine from i gram of 
opium extract. 

Example. — If 5.5 cc. of o.i n-potassium hydroxide solution were required to 
neutralize the excess of o.i n-hydrochloric acid in the 50 cc. of solution, then 
12.5 — 5.5 = 7 cc. of 0.1 n-hydrochloric acid are combined with morphine. Ac- 
cording to the proportion 

Cc. 0.1 n-HCl : Grams morphine 

1000 : 28.5 = 7 : X (x = 0.199s) 

7 cc. of O.I n-acid correspond to 0.1995 gram of morphine. Therefore this 
quantity of alkaloid is in i gram of extract. Consequently the opium extract 
contains 19.95 V^^ cent, of morphine. 

The German Pharmacopoeia requires that not more than 6.5 cc. nor less than 

5.5 cc. of 0.1 n-potassium hydroxide solution shall be used to produce a pale 
red color in the aqueous layer, corresponding to a morphine content of 17. 11 to 
19.95 per cent. 

3. In Wine of Opium and Tincture of Opium. — "Evaporate about 50 grams 
of either preparation in a weighed dish to 15 grams, add water until the weight 
is 38 grams and also 2 grams of sodium salicylate solution (1:2). Shake vig- 
orously and pass 32 grams of clear solution through a dry creased filter (10 cm. 
in diameter) into a dry flask. Mix this filtrate with 10 grams of ether by rotating 
the flask and add also 5 grams of a mixture of 17 grams of ammonium hydroxide 
solution and 83 grams of water." Continue the assay as directed above in i 
(Opium) from the point marked with an asterisk. 

Calculation. — Only 32 grams of the 40 grams of clear liquid (38 grams of evapo- 
rated opium tincture -f- 2 grams of sodium salicylate solution) are used for the 
morphine determination. These correspond to 40 grams of the original opium 
preparation. The morphine from this quantity of solution is dissolved in 25 
cc. of 0.1 n-hydrochloric acid and the volume brought to 100 cc. Excess of acid 
in 50 cc. of this solution is determined by titration. These 50 cc. contain the 
morphine from 20 grams of the opium preparation. 

Example. — If 4.2 cc. of 0.1 n-potassium hydroxide solution aite required for 



QnANTITATIVE ESTIMATION OF ALKALOIDS 



cc. of moiphine hydrochloride solution, then i 

ochloric add have combined with morphine. 

.t n<HCl : Grams morphine 



According to the proportion 



> 



(x = 0.2365s) 

ao grams of the opium prcparaCioii contain 0.13655 gram of morphine, corre- 
sponding to a morphine content of i.iS per cent. 

The German Pharmacopceia requires that not more than 5.5 cc. nor lesa than 
4.3 cc. of o.i n-potasslum hydroxide solution shall be used to produce a pale 
red color in the aqueous liquid, corresponding to a morphine content in Wine 
of Opium and Tincture of Opium of i.o to 1.18 per cent. 

Estimation of Pilocarpine in Jaborandum Leaves' 

G. Fromme's- Method.— Extract 15 grams of rather 
finely powdered jaborandum leaves with 150 grams of chloro- 
form and 15 grams of ammonium hydroxide solution (10 per 
cent. NHj), shaking frequently for 30 minutes. Filter this 
mixture through a large, smooth paper, covering the funnel 
with a glass plate. As soon as the chloroform drops slowly, add 
a little water and filtration will become more rapid. After 
collecting a full lOO grams of filtrate, add about i gram of water, 
shake vigorously and set aside. The water takes up fine 
particles of powder that may have passed through the paper, 
leaving the chloroform solution quite clean, After i hour weigh 
100 grams of chloroform solution (= alkaloids in 10 grains of 
jaborandum leaves). 

Fromme directs extracting these 100 grams of chloroform 
solution successively with 30, 20 and 10 cc. of i per cent, hydro- 
chloric acid which dissolves pilocarpine (and isopilocarpine) as 
hydrochlorides. Extract this acid solution first with 20 cc. of 
ether, to remove fat and resin. Then add an excess of ammonia 

id extract the free alkaloids successively with 30, 20 and 10 cc. 
chloroform. Pour the combined chiorofomi extracts through 
a dry iilter, evaporate in a weighed flask, dry the residue at 100° 
and weigh. 

2. Matthes and Rammstedt's* Method. — Evaporate 100 

> Further information about pilocarpine is given on page 21a. 
*C'aesBr and Loretz, Geschiftsbericht 1901, 37. 
■ See page 346. 



^etb 
Hani 



( 



280 DETECTION OF POISONS 

grams of chloroform solution obtained above in a beaker to 
about 20 cc. Add first 3 cc. of o.i n-picrolonic add and then 
60 cc. of ether. After 24 hours collect the precipitate of pilo- 
carpine picrolonate in a weighed Gooch crucible, wash with i cc. 
of alcohol-ether mixture (1:3), dry at no® and weigh. Pilo- 
carpine picrolonate thus obtained (= pilocarpine from 10 grams 
of jaborandum leaves), C11H16N2O2.C10H8N4O5 (Mol. Wt. 472) 
melts at 200-205°. 

Piperine in Pepper 

Black pepper is the dried, unripe fruit of the pepper plant, Piper nigrum L., 
whereas the ripe fruit deprived of its outer covering is the white pepper of com- 
merce. The actual constituents of pepper are piperine, an ethereal oil (oil of 
pepper) and a resin called chavicine. In rather large doses pepper is toxic. ^ 

Preparation of Piperine. — Extract finely divided white pepper with 90 per cent, 
alcohol and distil the latter from the extracts. Treat the residue with cold 
potassium hydroxide solution which dissolves the resin but not the piperine. 
Wash the residual piperine with water and crystallize from hot alcohol, using 
animal charcoal to remove color. White pepper contains 7-8 per cent, of piperine. 

Piperine, C17H19NOS, crystallizes in colorless, shining, rectangular, monocUnic 
prisms melting at 1 28-1 29°. When pure it is almost tasteless but impure piperine 
has a sharp, burning taste. It is nearly insoluble in water, freely soluble in alco- 
hol and also soluble in ether, benzene and chloroform. Piperine is a very weak 
base, dissolving in dilute mineral acids with almost as much difficulty as in pure 
water. A solution of piperine in concentrated sulphuric acid has a ruby color, 
soon changing to dark brown and gradually to greenish brown and fading upon 
addition of water. Concentrated nitric acid converts piperine into an orange 
red resin soluble with blood-red color in dilute potassium hydroxide solution. 
The constitution of piperine is known. 

Prolonged heating with alcoholic potassium hydroxide solution decomposes 
piperine into the potassium salt of piperic acid and piperidine: 

CH = CH.CO.NC»Hio CH = CH.COOK 

I -f KOH = C*HioNH + I 

CH = CH.C«H,(COaH,) Piperidine CH = CH.CeH,(CO,H0 

Piperine Potassium piperate 

Rtigheimer' synthesized piperine by putting together these two products. 
Piperic acid was first converted into its chloride by means of phosphorus penta- 

^ R. Robert ("Intoxikationen") mentions a case where a teaspoonful of 
pepper was given to each of three young pigs. There was severe inflammation 
of the gastro-intestinal tract in all three animals and two died. The toxic action 
of pepper is attributed to piperine, since the ethereal oil according to Kobert does 
not take part in the toxic action due to absorption. 

* Berichte der Deutschen chemischen Gesellschaft 15, 1390 (1882). 



QUANTITATIVE ESTIMATION OF ALKALOIDS 281 

chloride. Piperyl chloride was then condensed in benzene solution with piperi- 
dine: 

CH = CH.COOH CH = CH.COCl 

i +PCI» = I +P0C1, + HC1 

CH - CH.C,H,(CO,Ha) CH = CH.CeH,(CO,H,) 

CH = CH.C0C1 CH = CH.CO.NC»Hio 

I + HNC»Hio = I + HCl. 

CH = CH.CeH,(CO,H0 CH = CH.CeH,(CO,H,) 

Pix>eryl duoride Piperidine Pil>erine 

On the basis of the known structure of piperic acid and piperidine, piperine 
must have the following constitution: 

H H, 

C C 

^\ /\ 

yCh-C CH H,C CH, 

^O-C CH H,C CH, 

\/ \/ 

C N 

I 

— CH=CH.CH=CH.CO— I 

Estimation of Piperine in Pepper 

1. J. Koenig's Method. — Exhaust 10-20 grams of pepper, 
ground as finely as possible, in a Soxhlet apparatus with strong 
ethyl or methyl alcohol, or petroleum ether. Distil the alcohol 
or petroleum ether. The residue consists of piperine and resin. 
Shake this residue with cold potassium or sodium carbonate 
solution to dissolve the resin. Filter from undissolved piperine 
and wash the latter with cold water. Dissolve in alcohol or 
petroleum ether, evaporate the filtered solution in a weighed 
flask or dish and dry the residue at 100° to constant weight. 

To determine the resin in pepper at the same time, filter the 
potassium or sodium carbonate solution from crude piperine and 
add hydrochloric acid to the filtrate. Filter the precipitated 
resin, redissolve in alcohol, evaporate the solvent and dry the 
residue to constant weight. 

2. Cazeneuve and Caillot's Method. — Add enough water to 
make a thin mixture of powdered pepper with twice its weight 
of slaked lime and stir well. Boil in a porcelain dish, dry thor- 
oughly upon the water-bath and then extract with ether in a 
Soxhlet apparatus. Distil the ether in a weighed flask and 



282 DETECTION OF POISONS 

dry the residue of pipeline at loo® to constant weight. To 
obtain pure crystalline pipeline, dissolve the residue from the 
ether distillation in the least possible volume of boiling alcoh(d, 
surround the solution with ice, collect the piperine upon a 
weighed filter and dry at ioo° to constant weight. This 
purification of piperine is attended with more or less loss and 
consequently the result is only approximately correct. 

Estimation of Santonin in Wormseed^ 

I. K. Thaeter's^ Method. — Extract lo grams of crushed 
wormseed in a Soxhlet apparatus with ether for 12 hours. 
Distil the ether and boil the residue for an hour with 5 grams 
of lime and about 300 cc. of water. Replace water lost by 
evaporation. Filter while hot and wash the residue with 
water. Faintly acidify the filtrate with sulphuric acid and 
warm gently until santonin crystals begin to form. Then add 
100 grams of aluminium acetate solution,* heat the mixture 
to boiling and finally evaporate to dryness upon the water- 
bath. Mix the finely powdered residue with 3 grams of mag- 
nesium oxide, moisten this mixture with a little water and again 
bring quickly to dryness. Powder the residue as finely as pos- 
sible, dry at 105° and extract in a Soxhlet apparatus with an- 
hydrous, acid-free ether for 5 hours. Santonin is deposited 
upon distilling the ether as a faintly yellowish residue which 
is then dried at 100° to constant weight. 

Remarks. — When wormseed is heated with lime, santonin passes into solution 
as calcium santonate, and at the same time resinous substances are saponified. 

^ Wormseed (Flores dnae) consists of the unexpanded flower-heads of Artemisia 
cina which are 3-4 mm. in length. 

* Archiv der Pharmacie 237, 626-632 (1899) and 238, 383-387 (1900). 

' Dissolve 300 parts of aluminium sulphate in 800 parts of water; add acetic 
acid (sp. gr. 1.041) 360 parts; triturate caldum carbonate 130 parts with 200 
parts of water, and add this mixture slowly and with continued stirring to the 
first solution; set the whole aside for 24 hours without applying heat, and stir 
occasionally; then strain, press the precipitate without washing it and filter the 
liquid. It is a clear, colorless liquid, having the sp. gr. 1.044 to 1.046, a faint 
odor of acetic acid, an acid reaction, and a sweetish, astringent taste. National 
Dispensatory. 



QUANTITATIVE ESTIMATION OT ALKALOIDS 283 



I 



I 

I 



Dilute sulphuric arid liberates first santonic add which passes at once into its 
inner anhydride, santonin. Basic aluminium acetate, produced by iMiling, 
precipitates resinous imd colored substances. Finally, magnesium oxide serves 
to neutralize free acetic acid. Under the conditjuns, practically no magnesium 
santonate is formed. ThaeterobtaincdSS tog:i per cent, of the santonin present. 
Wormseed contains about a.j per cent, of santonin. 

2. J. Katz's' Method. — Extract lo grams of coarsely pow- 
dered wormseed ia a Soxhlet apparatus with ether for 2 hours. 
Distil the ether. There usually remains a dark, green resin 
weighing 1.5-2 grams. Boil this residue 15-30 minutes, under a 
reflux condenser, with 5 grams of crystallized barium hydroxide 
dissolved in 100 cc. of water. Cool and, without filtering, 
render the solution acid to litmus with carbon dioxide. Filter 
immediately with a pump to remove barium carbonate, and 
wash the precipitate twice with zo cc. portions of water. Evapo- 
rate the pale yellow solution to about 20 cc. in a dish upon the 
water-bath. Add 10 cc. of 12.5 per cent, hydrochloric add, 
continue heating upon the water-bath exactly 2 minutes longer 
and pour the solution into a separating funnel. Dissolve 
santonin crystals left in the dish in about 20 cc. of chloroform. 
Pour this solution into the separating funnel and shake thor- 
oughly. When the solutions have separated dear, withdraw 
the chloroform solution and pass it through a dry filter. Wash 
dish, separating funnel and filter 2-3 times with 10 cc. portions 
of chloroform. Distil the chloroform and boil the residue 10 
minutes, under a reflux condenser, with 50 cc, of 15 per cent, 
alcohol. Filter while hot into a weighed flask, and wash flask 
and filter twice with 15 cc. portions of 15 per cent, boiling 
alcohol. Cover the flask and set aside in the cold 24 hours. 
Weigh flask and contents and pass the latter through a weighed 
filter, disregarding the milky appearance of the filtrate caused by 
minute, resinous drops. Wash flask and filter once with 10 cc. 
of 15 per cent, alcohol, dry the filter in the flask and weigh both. 
Finally, apply a correction on account of the solubility of 
santonin in the alcohol used. Every 10 grams of filtrate contain 
6 mg, of santonin. Santonin by this method is crystalline, 

' Archiv der Pharmacie 337, 251 (iSpg). 



284 DETECTION OF POISONS 

and usually faintly yellow. J. Katz found the quantity of 
santonin in wormseed to vary between 1.2 1 and 3.16 (average 
2.42) per cent. This method is based upon the fact that the 
santonin in 10 grams of wormseed is easily soluble in 50 cc. of hot 
IS per cent, alcohol, whereas only a very little resin is dissolved 
by this dilute alcohol. As this dilute, alcoholic solution cools, 
santonin crystallizes out almost quantitatively. 

Troches of Santonin. — To estimate the quantity of santonin in troches, made 
from this substance and sugar, directly extract the finely ground mixture 
with hot chloroform. The santonin can usually be weighed without further 
purification. 

Chocolate Troches of Santonin.*— In a somewhat simpler form, the method 
described above may be used to estimate santonin in chocolate troches. Weigh 
3 or 4 troches and boil 15 minutes under a reflux condenser with 5 grams of barium 
hydroxide and 100 cc. of water. Saturate the liquid when cold with carbon 
dioxide. Filter, wash the residue with water and evaporate the brownish filtrate 
to 100 cc. Warm the liquid and add 10 cc. of dilute hydrochloiic acid. Three 
extractions with chlorcfcrm 3deld nearly pure santonin. To get santonin crys- 
tals almost white and ready for weighing, evaporate the chloroform solution and 
expel the last traces of chloroform by adding a few cc. of ether. If santonin is 
impure from traces of fatty acids, boil once with 10 cc. of petroleum ether and 
filter when cold. Santonin is nearly insoluble in cold petroleum ether. 

Santonin can be detected and estimated in toxicological analysis in a similar 
manner. Acidify the material with hydrochloric add, extract with chloroform 
and treat the chloroform residue with barium hydroxide solution as desaibed 
above. 

» 

Estimation of Solanine in Potatoes^ 

I. O. Schmiedeberg and G. Meyer's Method. — Mix 500 

grams of finely grated potatoes with water and press out the 
liquid. Decant the liquid from the deposit of starch. Again 
mix the starch with water and decant the latter when the 
starch has settled. Neutralize the entire liquid with ammonia 
and evaporate to the consistency of an extract. In the mean- 
time mix the press cake with several times its volume of boiling 
alcohol. Press out the alcohol completely after several hours. 
Make two such extractions. Filter the combined alcoholic 
extracts and wash the residue (starch) upon the filter with 
alcohol. The aqueous liquid from the potatoes contains very 

* See page 217. 



QUANTITATIVE ESTIMATION OF ALKALOIDS 



285 



little solatiine. To isolate this small quantity, use the alcoholic 
filtrate to extract the residue from the aqueous extract and 
again filter. Wash the insoluble part with hot alcohol. The 
alcoholic filtrate after half an hour usually deposits some 
crystals of asparagine.^ Separate the supernatant liquid from 
these crystals and evaporate upon the water bath to the con- 
sistency of an extract. Dissolve the residue in water con- 
taining sulphuric add, filter and wash. Warm the clear 
liquid very gently, saturate with ammonia and set aside for a 
day. Solanine appears in small crystals. Collect the deposit 
upon a weighed filter, wash first with water and then with ether, 
dry at 100° and weigh. 

2. F. von Morgenstem's' Method. — Express as much 
liquid as possible from 200 grams of finely grated potatoes by 
by means of a press. Make two separate extractions of the 
press cake with water and express the liquid thoroughly each 
time. Precipitate protein sustances from the combined liquid 
by adding 0.5 cc. of acetic acid and warming for i hour upon the 
water bath. Filter, evaporate the filtrate to a syrup, stir and 
add gradually hot 96 per cent, alcohol until cloudiness ceases.* 
Decant the solution after 12 hours and extract the residue 
containing sugars and dcxtrins twice with hot alcohol. Evapo- 
rate the combined alcoholic extracts upon the water-bath, warm 
the residue with some water containing acetic acid and lilter. 
Heat the filtrate to boifing and add ammonia drop by drop to 
precipitate solanine. After standing for 5 minutes upon the 
water-bath the base separates in flocks that are easily filtered. 
Wash the precipitate with water containing ammonia, dissolve 
in boiling alcohol and treat this solution as follows. Evaporate 

' Asparagine is Ihe amide of ospattic acid, or mono- amino-succinic &dd, 
HJ^.CH-COOH 

I -|- HiO. It appears in shining, rhombic crystals that 

CH,-CO.NH, 
dissolve rather easily in hot water but less easily in alcohol or ether. La;vD- 
BSpaiagine is widespread in the plant kingdom in seeds. 
' Landwirtschaftliche Versuchsstation 65, joi (1507). 

' To eitfact those parts of the potato plant, which can be dried at 100° and 
reduced to a fine powdei, heat lo boiling several times with water containing 
c add and filler each lime. 



286 DETECTION OF POISONS 

upon the water-bath and dissolve the residue in water containing 
acetic acid. Filter, heat the filtrate to boiling and precipitate 
solanine with ammonia. Collect the pure white flocks of sola- 
nine from this second precipitation upon a filter that has been 
dried at 90° and weighed. Wash with 2 per cent, ammonia and 
dry at 90° to constant weight. 

Notes.i— V. Moigenstern obtained on the average by this method 0.0135 per 
cent, of solanine in table potatoes and 0.0058 per cent, in those used as forage. 
The yield of solanine fi om yellow tubers upon the average was less than from red. 
Tubers grown upon sandy soil were richer in solanine than were those from humus 
soil. Moistuie and abundance of humus appear to diminish the quantity of 
solanine. A nitrogenous fertilizer increased the quantity of solanine, a potash 
fertilizer lowered it and a phosphate fertilizer appeared to have little effect. 
There was less solanine in large than in small potatoes of the same vaiiety. 
Solanine first appears to increase during the process of germination. Passing 
into sprouts, without wholly disappearing from the tubeis, solanine increases 
with the growth of the plant. As growth advances the distribution of solanine 
in the different parts of the plant is indication of a tendency on the part of the 
plant to withdraw solanine from the older sprouts and spread it throughout the 
young organs. Consequently solanine may serve first of all as the natural 
protector of the plant, especially of the growing parts. 

Estimation of Alkaloids in Nux Vomica 
(C. C. KeUerO 

Remove fat from mix vomica by treating 15 grams of the 
well-dried and finely powdered drug in a 250 cc. Erlenmeyer 
flask two or three times with 30 cc. portions of ether. Shake 
thoroughly for 5 minutes. Pour these ether washings into a 
flask, and, since they contain a little alkaloid, extract dissolved 
alkaloid with 5 cc. of o.i n-hydrochloric acid and 10 cc. of water. 
Repeat the extraction of the ether layer, separated from the 
aqueous solution, using water instead of acid. Add 100 cc. of 
ether, 50 grams of chloroform and 10 grams of 10 per cent, am- 
monium hydroxide solution to the powdered nux vomica free 
from fat. Shake thoroughly for 30 minutes and add to this 
mixture the hydrochloric acid solution used in extracting alka- 

' Festschrift presented at the fiftieth anniversary of the founding of the Swiss 
Pharmaceutical Association. Abstract in 2^itschrift fUr analytische Chemie, 
23, 491 (1894). 



r 



QUAKTITATIVE ESTIMATION OF ALKALOIDS 287 

loid from the first ether washings. Again shake thoroughly 
and, when the liquids have separated clear, pour loo grams of 
ether-chloroform solution through a small filter into a weighed 
Erlenmeyer flask. Distil the chloroform and ether as com- 
pletely as possible. The alkaloids usually appear as colorless 
» varnishes which persistently retain chloroform. To remove the 
latter, pour a few cc. of absolute alcohol upon the residue and 
ejtpel completely upon the water-bath. Repeat this treatment 
S-3 times. This will give crystalline alkaloids which can be dried 
at iDO° to constant weight. 



Method of the German Phaimacopoeia 



In Nui Vomicft. — Place is grams oi nux vomica, ground mediumly fine 
ftnd diied at 100°, in an Erlenmeyer flask and add 100 grams Qf ether and 50 
grains of chloroform. Shake vigorouEly and add xo cc. of a mixture of z parts 
of sodium hydronide solution and t part of water. Shake at frequent intervals 
•nd lei the mixture stand (oi 3 hours. Then add 1 s cc. more of watei , or enough 
to cause thepowdcrafter vigorous shaking to gather into balls and leave Ihesupet- 
natant ethei -chloroform solution peifectly clcai. After i houi filter loo grams 
o£ the cleat ether -chloroform solution through a dry filter kept will covered. 
Collect the filtrate in a small flask and distil about half the solvent. Transfer 
the residual elher-cblorpfotm solution to a separating funnel, riose the flask 
3 times with 5 cc. portions of a mixture of 3 parts of ether and 1 port of chloroform. 
Extract the combined solvent with 10 cr. of o. 1 n- hydrochloric add. Add enough 
ether to cause the cther-chlorofoim solution to rise to the lop of the add liquid 
and pass the lattei through a small filter moistened with water into a loo cc. 
flask. Then extract the ether-chloroform solution with 3 additional 10 cc. por- 
tions of water. Pass these extracts through the same filter, wash the lattei with 
watei and dilute the total liquid to 100 cc. Finally measure 30 cc. of this solution 
into a flask holding about 300 cc, add about jo cc, of water and sufficient ether 
to make a laj'Ct i cm. deep. Add 5 drops of iodeosine solution and run in enough 
o.oi n-potassium hydroxide solution, shaking vigorou°ly after each addition, to 
turn the aqueous Inyei a permanent pale red. 

CalciiUtion.— 100 grams {= alkaloids from 10 grams of nux vomica) of ihe 
original 150 grams of ether-chloroform mixture were used. The alkaloids were 
dissolved by 10 cc. of o.i n-hydrochloiic add and the volume was brought to 
100 cc. The excess of add in 50 cc. of this solution ( = s grams of nux vomica) 
was determined by titration with o.oi n-potassium hydroxide. I( strychnine 
and brucine are present in nux vomica ill equal amount, the average equivalent 
weight of the two alkaloids is 364. Therefore 1000 cc. of o.i n-hydrochloric 
add correspond to 36.4 grams of alkaloids. 

Example. — Suppose that the titration of the excess of add In 50 cc, of solution 
required ts.6cc. of 0,01 n-potassium hydroxide = j.s6cc.of 0.1 n-alkali. Then 



288 DETECTION OF POISONS 

5 — 1.56 » 3.44 cc of o.z n-hydrochloric add are combined with the alkaloidi 
in 5 grams of nuz vomica. According to the proportion 

Cc. 0.1 n-HCl: Grams alkaloid 

1000 : 36.4 » 3.44 : X (x B 0.12533) 

3.44 cc. of 0.1 n-add are combined with 0.12522 gram of alkaloid, correspond- 
ing to an alkaloid content of 20 X 0.12522 » 3.50 per cent. The German 
Pharmacopoeia places this percentage as the minimum for total alkaloids in 
nuz vomica. 

a. In Extract of Nuz Vomica. — Dissolve i gram of extract in an Erlenmeyer 
flask in 5 grams of water and 5 grams of absolute alcohol, and add 50 grams of 
ether and 20 grams of chloroform to this solution. Shake vigorously and add 10 
cc. of sodium carbonate solution (i :$), Let the mixture stand and agitate at 
frequent intervals for an hour. * Then pass 50 grams of the dear ether-chloroform 
solution through a dry filter kept well covered, and recdve the filtrate in a flask. 
Distil half the solvent and pour the remainder into a separating f imnel. Wash 
the flask three times with 5 cc. portions of a mixture of 3 parts of ether and 
I part of chloroform. Thoroughly extract the total ether-chloroform solution 
with 50 cc. of o.oi n-hydrochloric add. When the liquids have separated 
dear, if necessary, after addition of enough ether to bring the ether-chloro- 
form solution to the surface, pass the add solution through a smaU filter 
moistened with water and recdve the filtrate in a 200 cc. flask. Make three 
extractions of the ether-chloroform solution with 10 cc. portions of water, and 
pass these washings through the same filter. Finally, wash the filter with water 
and bring the entire solution to 100 cc. Add enough ether to make a layer i cm. 
thick and 5 drops of iodeosine solution. Run in o.oi n-potassium hydroxide 
solution, shaking vigorously after each addition, until the aqueous solution is 
pale red. 

Calculation. — Only 50 grams, or two-thirds of the original 75 grams of alcohol- 
ether-chloroform mixture, were used. The alkaloids in 0.666 gram of nux 
vomica extract were in this volume of solvent. Alkaloids were dissolved in 50 
cc. of O.OI n-hydrochloiic add and excess of add determined by titration with 
O.OI n-potassium hydroxide solution. 

Example. — Suppose that 18 cc. of o.oi n-potassium hydroxide solution were 
used in this titration. Then 50 — 18 = 32 cc. of o.oi n-add were combined 
with the alkaloids in 0.666 gram of extract. According to the proportion 

Cc. o.oi n-HCl : Grams alkaloids 

1000 : 3.64 = 32 : X (x = 0.1 1648) 

0.666 gram of extract contains o. 11 648 gram of alkaloids, corresponding to 17.47 
per cent. The German Pharmacopoeia places this percentage as the minimum 
for total alkaloids in extract of nux vomica. 

3* In Tincture of Nux Vomica. — Evaporate 50 giams of tincture of nux vomica 
in a weighed dish to 10 grams. Wash this residue into an Erlenmeyer flask and 
rinse with 5 grains of absolute alcohol. Add 50 grams of ether and 20 grains of 
chloroform and shake vigorously. Then add 10 cc. of sodium carbonate solution 
(1:3) which has been previously employed in linsing the dish used in evaporating 
the tincture. Let this mixture stand an hour, shaking vigourously at fre- 



QUANTITATIVE ESTIMATION OF ALKALOIDS 289 

quent intervals. Filter 50 grains of the clear ether-chloroform solution. To ex- 
tract alkaloids, use 40 cc. of o.oi n-hydrochloric add. In other respects, the 
estimation of alkaloids is the same as described for extiact of nux vomica. 

CalcolatiQn. — This is the same as that given for extract of nux vomica. Only 
two-thirds (= 33.3 grams) of the original weight of nux vomica tincture were 
used. The alkaloids were dissolved in 40 cc. of o.oi n-hydrochloric acid and ex- 
cess of acid was determined by titration with o.oi n-potassium hydroxide 
solution. If 17 cc. of the latter solution were used, then 40 — 17 = 23 cc. of 
O.OI n-hydrochloric acid were combined with the alkaloids in 33.3 grams of the 
tincture. According to the proportion 

Cco.oi n-HCl : Grams alkaloids 

1000 : 3.64 = 23 : X (x = 0.08372) 

this weight of tincture contains 0.08372 gram of alkaloids, corresponding to 2.51 
per cent. The German Pharmacopoeia places this percentage as the minimum 
for total alkaloids in tincture of nux vomica. Brucine and strychnine are assumed 
to be present in equal quantity. * 

Estimation of Alkaloids in Nux Vomica and Its Preparations by Means 

of Picrolonic Acid 

(H. Matthes and O. Rammstedt) 

Nux vomica upon the average contains strychnine and 
brucine in equal quantity combined with tannic acid. Alkaline 
hydroxide or carbonate solutions liberate the alkaloids from 
their salts. The free bases are then extracted with an ether- 
chloroform mixture. The solvent is reduced to smaller volume 
by evaporation or distillation and the alkaloids are precipitated 
with picrolonic acid. 

Strychnine picrolonate, C21H22N2O2.C10H8N4O6 (Mol. Wt. 
S98) melts with decomposition at 286°. 

Brucine picrolonate, C23H26N2O4.C10H8N4O6 (Mol. Wt. 658) 
melts with decomposition at 277°. 

I. Extract of Nux Vomica. — Dissolve i gram of extract in 5 
grams of absolute alcohol and 5 grams of water. Shake well 
with 50 grams of ether and 20 grams of chloroform. Add 10 cc 
of sodium carbonate solution (i 12) and again shake thoroughly 
for 10 minutes. Let the mixture stand at rest for 20 minutes. 
Pass 50 grams of the ether-chloroform mixture through a dry, 
double, creased filter and evaporate half the solvent in a beaker. 
Add about 5 cc. of o. i n-alcoholic picrolonic acid to the warm 

19 



290 DETECTION OF POISONS 

solution. A yellow crystalline precipitate of strychnine and 
brucine picrolonates soon appears. After 24 hours collect the 
mixed picrolonates in a weighed Gooch crucible. Wash 
excess of picrolonic acid from the precipitate with 2 cc. of an 
alcohol-ether mixture (i 13), dry 30 minutes at no®, cool in 
desiccator and weigh. 

CalcttUtion. — Use the mean molecular weight of brudne and strychnine 
picrolonatesC — 628) and also the mean molecular weight of brudne and strych- 
nine ( — 364). The proportion is 

Grams picrolonate Grams strychnine Wt. of pre- 
mixture : and brudne « dpitate : x. 

628 : 364 

Since the quotient 364 '.628 >- 0.5798, the weight of mixed alkaloids is obtained 
by multiplying the weight of the picrolonate precipitate by this quantity. This 
predpitate represents total alkaloids in 0.666 gram of extract of nuz vomica, 
for only two-thirds ( » 50 grams) of the original 75 grams (5 grams of alcohol + 
50 grams of ether + 20 grams of chloroform) of solvent, containing the alkaloids 
in I gram of extract of nux vomica, were used. 

2. Tincture of Nux Vomica. — Evaporate 50 grams of tincture 
in an Erlemneyer flask to 10 cc. Cool and shake well with 5 cc. 
of absolute alcohol, 50 grams of ether and 20 grams of chloro- 
form. Add 10 cc. of sodium carbonate solution (i : 2) and shake 
again for 10 minutes. After 20 minutes pass 50 grams 
(= two- thirds of the original mixture) of the ether-chloroform 
mixture through a double, creased filter. Evaporate half the 
solvent in a beaker and add 5 cc. of o.i n-alcoholic picrolonic 
acid to the warm residue. Treat the picrolonate precipitate as 
described above. 

3. Nuz Vomica. — Exhaust 15 grams of powdered nux vomica, 
previously dried at 100, by thoroughly agitating with 100 grams 
of ether and 50 grams of chloroform. Then add 10 cc. of a 
mixture of 2 parts of 15 per cent, sodium hydroxide solution and 
I part of water and shake again for 10 minutes. Add an ad- 
ditional IS cc. of water, or enough to cause the powder to gather 
into balls after vigorous agitation and leave the supernatant 
ether-chloroform mixture dear. After 30 minutes pass the 
dear ether-chloroform solution through a dry, double, creased 
filter. Evaporate 50 cc. of the filtrate in a beaker nearly to 



QUANTITATnrE ESTIMATION OF ALKALOIDS 



291 



I 



dryness and add a second 50 cc. portion of filtrate to the 
idue, bringing everything into solution ( = alkaloids from 10 
grams of nux vomica). Add 5 cc. of o. i n-a!cohohc picrolonic 
acid and treat the precipitate as previously described. 

Eadmadon of Strychnine in Mixtures of Nux Vomica Alkaloids 

(Gordin's' ModificaLion of Keller's Method) 
Strong nitric acid, gently heated with a solution of strych- 
nine and brucine in 3 per cent, sulphuric acid, is without action 
upon the former alkaloid. But brucine is converted into non- 
basic substances not extracted by chloroform from an alkaline 
solution. 

Procedure.^Dissolve the mixed alkaloids (0.2 — 0.3 gram) 

upon the water-bath in 15 cc. of 3 per cent, sulphuric acid and 

badd 3 cc. of a diluted nitric add (equal parts of 68-69 P^'' cent. 

I add (sp. gr, 1.43) and water) to the cold solution. Pour the 

' mixture into a separating funnel after exactly 10 minutes and 

add sodium hydroxide solution in excess. Extract strychnine 

3-4 times with chloroform. Pass the chloroform solution 

through a double filter into a small weighed flask, wash the 

filter with a little chloroform, add 2 cc. of pure amyl alcohol and 

distil to dryness upon the water-bath. Remove the last traces 

of liquid, consisting mainly of amyl alcohol , by forcing a current 

of air through the flask warmed upon the water-bath. Finally 

dry the residue 2 hours at 135-140° and weigh when cold. In 

this way a very pure, white strychnine free from brudne is 

obtained. 

Hotes.^ — According lo Gordtn, ammoma cannot be substituted for sodium 
hydroxide, for it gives colored strychnine. Amyl alcohol is added to the chloro- 
form solution to prevent strychiune crystals from being carried by deciepitEitioD 
into the condenser during distillation. 

Estiination of Theobromine and Caffeine in Cacao and Chocolate' 

Cacao and its preparations contain only very little caffeine 
which is usually determined with theobromine. 

' Archiv der Pharmaiie 540, 643 (190J). 
H. Beckurts, Archiv der Phirmiiie, 244, 486 (1906). 



292 DETECTION OP 

Bofl 6 grams of powdered cacao, or 12 grams of diocolate, for 
30 minutes under a reflux condenser in a weired liter flask with 
200 grams of a mixture of 197 grams oi water and 3 grams of 
dilute sulphuric add. Then add 400 grams of water and 8 
grams of finely powdered magnesia and IxmI for an hour kmger. 
When the mixture is cold, add exactly oiou^ water by wei^t 
to replace what has been lost by ev2qx>ration. After the mix- 
ture has settled, filter 500 grams of solution (= 5gramsof cacao 
or ID grams of choccdate) and evaporate the filtrate to dryness 
in a dish either by itself or with some quartz sand. Triturate 
the residue and exhaust in a Soxhlet tube with chlorofonn. 

In case of evaporation without quartz sand, rub the residue 
with a few droi>3 of water, transfer to a separating funnel with 
ID cc. of water and extract 8 times with 50 cc. portions of hot 
chloroform. Pass the chloroform extract through a dry filter 
into a tared flask, distil the chloroform and dry the residue 
(= theobromine and caffeine) at 100^ to constant wdghL 

Carbon tetrachloride is used to separate theobromine from 
caffeine, the latter alkaloid alone being soluble at room tem- 
perature. Let the weighed residue from chloroform stand for i 
hour with 100 grams of carbon tetrachloride at room tem- 
perature. Shake occasionally and then filter. Distil carbon 
tetrachloride and extract the residue repeatedly with water. 
Evaporate the aqueous solution in a weighed dish and dry the 
residue (= caffeine) at 100^ to constant weight. 

Repeatedly extract the theobromine, insoluble in carbon tetra- 
chloride, and also the filter paper with water. Filter, evaporate 
the total filtrate and weigh the residue ( = theobromine) dried 
at 100°. 

Notes. — In the method described above, H. Beckurts and Fromme rfiminate 
injurious enects due to concentration by boiling with dilute sulphuric mcid. 
Xanthine bases are set tree from combination with organic acid and reoooibined 
with sulphuric add. Magnesia sets these bases free from their sulphates and 
at the same time holds back cx^loiing matter and £at, thus cfiminadng these 
impurities. 

Theobromine^ 3. 7>dime:hyl- xanthine, CtH^XiOs. is a white powder ooosisdng 
of microscopic needles haxin^: a bitter taste. It dissolves in 3282 parts of cold 
and 148 paru of boiling water; in 422 parts of boiling absolute alcohol; and in 



QUANT1TATI\-E ESTIMATION OF ALKALOIDS 



293 



I 



105 parts o( boiling chloroform. Theobromiiie solutions are neutral. Tbis 
alkaloid acts both as an acid and as a base and therefore is soluble in b«th add 
and alkaline solutions. The salts with acids crystalline well but are not very 
stable. These theobromine salts are partially decomposed into theobromine 
and acid in presence of much water, or. if the given acid is volatile, by heating at 
100°. Theobromine is isomeric with theophylline, or 1,3 -dime thy l-xanthine, 
kDd paraxanlhinc, or 1,7-diincthyl-xanthine: 

HN— CO (i)CH..N— CO (i)CH,.N— CO 

I I /CH.(7) i i H 1 I /CH.(7> 

DC C— N< OC C— N^ OC C— N< 

t J! >CH I II >CH I ]| >CH 

(3)CH,.N-C— N'*^ (3)CH,.N-C— N**^ HN=C— N^ 

Theobromins Thoophylline Paruanthine 

Theophylline occurs in tea leaves and paraxan thine has been isolated fiom human 
mine. The latter is therefore called urotheobromine. 



Ukaloids in Leaves of Atropa Belladonna, HyoscyamuB 
I Niger and Datura Strammonitun 

(E. Schmidt's Modification of Keller's Method') 

Shake vigorously 10 grams of finely powdered leaves, dried to constant weight 
over quicklime, in an Erlenmeyer Bask, with 90 grams of ether and 30 grams of 
chloroform. Add 10 cc. of 10 per cent sodium hydroxide solution and shake 
vigorously and often for 3 hours. Then add 10 cc. of water, or enough to cause 
the powder to gather into balls when thoroughly shaken. After 1 hour pass 60 
grams of the ether-chloroform extract (= 5 grams of leaves) through a di; 
filter kept well covered. Distil 60 cc. of this filtrate to half its volutne to remove 
ammonia, and transfer the deep green solution to a separating funnel, rinsing 
the fiask with three j cc. portions of ether. Shake the combined extracts well 
with 10 cc. of o.oi n-hydrochloric acid. Add enough ether to cause the ether- 
chloroform solution to rise to the top, and pass the acid solution through a moist 
filter into a 100 cc. glass stoppered flask. Shake the ether-chloroform solution 
3 times with 10 cc. portions of water, pouring these extracts through the same 
filter and washing the latter with enough water to bring the total volume to too 
cc. Add enough ether to make a layer 1 cm. deep and 5 drops of iodcosine solu- 
tion. Having determined befcrehond the exact relation of add to alkali, titrate 
excess of o.oi n-hydrochloric add with 0.01 n-potassium hydroxide solution. 
The calculation is the same as that for extract of belladonna (see page igt). 

Notes. — U^ng ihis method, E. Schmidt obtained 0.4 per cent, of alkaloid in 
wild belladonna leaves but only 0.16 per cent, in cultivated leaves. The average 
of many determinations gave 0.4 per cent, instrammonium leaves ando.jT-^.aS 
per cent, in hyoscyamus leaves without stalks. Alkaloids were calculated at 

Sodium hydroxide solution liberates alkaloids from the odds with which they 
re naturally combined in the plant, for example: 

(C,jH„NO,),.H,SO,' + jNaOH - jChHuNOi + iH.Q -t- Na,SO,. 

' Apotheker-Zeitung 15, 13. 

' The formula of atropine sulphate used in medidne. 



294 DETECTION OF POISONS 

Estimatioii of Alkaloids in Extract of Belladonna 

(German Pharmacopoeia) 

Dissolve 2 grams of extract of belladomia in an Erlenmeyer flask in 5 grams of 
water and 5 grams of absolute alcohol, and add 50 grams of ether and 20 grams of 
chloroform to this solution. Shake vigorously and add 10 cc. of sodium carbonate 
solution (i 13). Let the mixture stand and agitate at frequent intervals for an 
hour. Then pass 50 grams of the clear ether-chloroform solution through a dry 
filter kept well covered, and receive the filtrate in a flask. Distil half the solvent 
and pour the remainder into a separating funneL Wash the flask three times 
with 5 cc. portions of ether. Thoroughly extract the total ether-chloroform so- 
lution with 20 cc. of o.oi n-hydrochloric add. When the liquids have separated 
dear, if necessary, after addition of enough ether to bring the ether-chloro- 
form solution to the surface, pass the add solution through a small filter moist- 
ened with water and receive the filtrate in a 200 cc. flask. !Make three extrac- 
tions of the ether-chloroform solution with 10 cc. portions of water, and pass these 
washings through the same filter. Finally, wash the filter with water and bring 
the entire solution to 100 cc. Add enough ether to make a layer i cm. thick 
and 5 drops of iodeosine solution. Run in o.oi n-potassium hydroxide solution, 
shaking vigorously after each addition, until the aqueous solution is pale red. 

Calculation. — Sodium carbonate Uke sodium hydroxide liberates the alkaloids 
atropine and hyoscyamine, from their salts in belladonna leaves: 

(C,7H«NO,)j.H,S04 -f Na,CO, = 2Ci7H„NOi -f Na,S04 + CO, + H,0. 

The free alkaloids dissolve in the alcohol-ether-chloroform mixture. Fifty grams 
of this solution ( >■ alkaloids from 1.33 grams of extract) are extracted with 20 cc. 
of O.OI n-hydrochloric add, the alkaloids passing into the aqueous solution as 
salts of hydrochloric add (CnHssNOs.HCl). Excess of acid in this solution is 
determined by titration. If 13 cc. of o.oi n-potassium hydroxide solution are 
used, then 20-13 » 7 cc. of o.oi n-hydrochloric add correspond to the alkaloids 
in 1.33 grams of extract. The equivalent weight of the two isomeric bases, 
atropine and hyoscyamine, being 289, 1000 cc. of o.oi n-hydrochloric add corre- 
spond to 2.89 grams of alkaloids. The proportion 

1000 : 2.89 = 7 : X (x = 0.02023) 

shows that 1.33 grams of belladonna extract contain 0.02023 gram of alkaloids 
corresponding to 1.51 per cent. The German Pharmacopoeia places this per- 
centage as the minimum for total alkaloids in extract of belladonna. 

Extract of Hyoscyamus 

The alkaloids in 2 grams of this extract are determined in the manner described 
for extract of belladonna. Use 10 cc. of o.i n-hydrochloric acid instead of 20 cc. 
to extract alkaloids. The German Pharmacopoeia requires that not more than 
6.5 cc. of O.OI n-potassium hydroxide solution shall be used in titrating the excess 
of hydrochloric add. Therefore 10—6.5 = 3-5 cc. of o.oi n-hydrochloric acid 
are combined with the alkaloids in 1.3 grams of henbane extract (= two- thirds 
of the original extract). The proportion 

1000:2.89 ^ s,s:x (x = o.oioii) 



QUANTITATIVi; ESTIMATION OF ALKALOIDS 



diows that 1-33 giatas of extract contain o.oi 
to 0.76 per cent. This percentage is placed a) 
henbane extract. 



I 



Assaying Officinal Extracts 

(E. Merck") 



,s possible, E, Merck has 



With n view to obviating as many sources 
proposed the folloning procedures: 

Extract of Belladonna.— Dissolve 4 grams of extract in 6 cc. of water and wash 
the solution into a separating funnel with an additional 10 cc. Add 100 cc. of 
ether, shaking well, then 10 cc. of sodium carbonate solution Ti :3) and shake at 
once [or s minutes. Stopper the funnel and let the mixture stand for to minutes. 
Then pass the ether layer through a dry filter (10 cm. in diameter) into a gtass- 
stoppered flask. To lessen evaporation of ether as much as possible, cover the 
funnel with a glass plate. If an emulsion keeps the ether from separating well, 
add a few grams of powdered tragacantb at the end of the time stated above. 
Shake until the tragacanth gathers into balls in the aqueous layer. After is 
minutes decant 75 cc. of the ether layer. To check results by making more than 
one assay, use 7$ cc. of the ether solution (^^ i gram of extract). Test a clean 
glass-stoppered Sask to make sure that it docs not give up alkali to the water. 
Then introduce into such a flask 50-60 cc. of water, 5 drops of iodeosine solution 
and 20 cc. of ether. Shake and add o.oi n-hydrochloric acid until the aqueous 
layer just becomes colorless upon shaking. This procedure obviates a special 
determination of the alkalinity of the water, since the resulting mixture is brought 
to the neutral point. Now add 35 cc. of the ether solution of the alkaloid and 
titrate until there is no color. M ultiply the number of cc. of o.oi n-hydrochloric 
add used by o.ooiSg.* The product is the quantity of alkaloid, calculated as 
atropine, in 1 gram of belladonna extract. Upon the average this preparation 
contains i.S per cent, of alkaloid. 

Bxtroct of Cinchona. — HsmatoxyUn is frequently an unsatisfactory indicator 
in the titration of cinchona alkaloids, because the color change is slow enough 
to make it difljcult to fix the end point exactly. Therefore E, Merck makes a 
gravimetric and volumetric determination at the same time by the following 
method: 

Dissolve 3 grams of aqueous cinchona extract in lo cc. of water in a porcelain 
dish. Pour the solution into a 350 cc. shaking flask, rinsing it in with 10 cc. 
of water. Add 150 cc. of ether and 10 cc. of sodium carbonate solution (i :i) 
to this mixture and shake vigorously for 10 minutes. Cork the flask and let the 
mixture stand at rest for 30 minutes. This extract frequently forms an emulsion. 
In that case add a few grams of tragacanth powder which has no effect upon the 
result. Pour the ether solution of cinchona alkaloids as rapidly as possible 
through a dry creased filter. Use 50 cc. ( = 1 gram of cinchona extract) for each 

■ Zeitschrift fiir analytische Chemie 41, 584 (1903) and also Merck's Bericht 
Qber das Jahr 1900, 

• 189 = the equivalent weight of the two isomeric bases, atropine and hyosey- 
unine, CnHuNO). 



296 DETECTION OF POISONS 

determination. Distil the solvent from the 50 cc. in a weighed zoo cc flask and 
dry the residue in an air bath at loo-iio® to constant weight. The alkaloids 
obtained are nearly colorless or faintly yellow. Having ascertained the weight 
of the alkaloids, proceed with the titration. Dissolve the residue in the flask in 
10 cc. of alcohol, adding 50 cc. of water, which partially precipitates the alkaloids, 
and then alcoholic haematoxylin solution.^ Run in o.i n-hydrochloric add until 
the alkaloids. again dissolve and the red color of the solution passes through 
reddish yellow into a pure yellow. The mean equivalent weight of the cinchona 
alkaloids is 309. Therefore i cc. of o.i n-hydrochloric add » 0.0309 gram of 
alkaloid. Upon the average, officinal aqueous extract of cinchona contains 9 
per cent, of alkaloid. 

Extract of Nux Vomica. — Dissolve 0.1 gram of this extract in a flask in 5 
grams of absolute alcohol and 10 grams of water. Add 95 grams of ether and 
shake well. Then add 10 cc. of sodium carbonate solution (i :3) and shake 
vigorously at once for about 10 minutes. After 15 minutes poiu: the ether solu- 
tion as rapidly as possible through a creased filter. Weigh in a flask 50 grams of 
this solution (— 0.05 gram of the original extract), having previously placed in 
this flask a neutral mixture of 50 cc. of water, 20 cc. of ether and 5 drops of iod- 
eosine solution. Add 20 cc. of o.oi n-hydrochloric add and titrate with o.oi 
n-potassium hydroxide solution until the aqueous layer is just red. 

Calculation. — Since strychnine and brucine are present in nux vomica in nearly 
equal parts, the mean equivalent weight of such a mixture of bases (334 + 394) : 3 
■■ 364. Hence 0.00364 gram of the mixed alkaloids neutralizes i cc. of o.oi 
n-hydrochloric add. The officinal extract of nux vomina contains 18 per cent, 
of alkaloid. 

^ £. Merck advises keeping on hand an alcoholic solution of haematoxylin, 
because a freshly prepared solution usually gives a blue-violet instead of a red 
color change. 



CHAPTER VII 

DETECTION OF CARBON MONOXIDE BLOOD, BLOOD STAINS 

AND HUMAN BLOOD 

I. Carbon Monoxide Blood 

Carbon monoxide (CO) has a direct toxic action upon the 
blood. This gas passed into blood displaces loosely bound 
oxygen from oxyhaemoglobin forming the more stable carboxy- 
haemoglobin. The latter compound is cherry red, not dichroic 
and entirely resistant to putrefaction if air is excluded. In 
carbon monoxide poisoning the cherry red color of the blood 
is usually noticed at once. 

Detection of Carbon Monoxide Blood 

1. Boiling Test. — Blood containing carbon monoxide gives a 
brick red coagulum, if boiled or warmed upon the water-bath. 
Ordinary blood gives a grayish brown or brownish black 
precipitate. 

2. Sodium Hydroxide Test — Carbon monoxide blood shaken 
with 1-2 volumes of sodium hydroxide solution (sp. gr. 1.3 = 26.8 
per cent.) remains red and in a thin layer is the color of red lead 
or vermilion. Normal blood similarly treated is almost black 
and in a thin coating upon a porcelain plate is dark greenish 
brown. A procedure recommended consists in diluting the 
blood with 6-10 times its volume of water and using about 5 
drops of sodium hydroxide solution to 10 cc. of diluted blood. 
Even gentle warming with sodium hydroxide solution do per 
cent. NaOH) does not alter the red color of this carboxyhaemo- 
globin solution, whereas a solution of normal human blood 
becomes greenish to dark brown. 

3. Basic Lead Acetate Test — Mix 4-5 volumes of basic 
lead acetate solution in a test-tube with diluted or undiluted 
carbon monoxide blood and shake vigorously for a minute. 

297 



298 DETECTION OF POISONS 

Such blood remains bright red but normal blood is first brown- 
ish and then chocolate to greenish brown. 

4« Potassium Ferrocyanide Test — Mix undiluted blood (15 
cc.) with an equal volume of 20 per cent, potassium ferrocyanide 
solution and 2 cc. of diluted acetic acid.^ Shake the mixture 
gently and a coagulum will gradually form. That from normal 
blood is dark brown but from blood containing carbon monoxide 
bright red. This difference disappears slowly but not entirely 
for weeks. 

5. Tannin Test — Mix an aqueous blood solution* with 3 times 
its volume of i per cent, tannin solution and shake thor- 
oughly. A difference in color between normal and carbon 
monoxide blood can be recognized after several hours, most 
distinctly after 24 hours. Normal blood is gray but carbon 
monoxide blood is crimson red. This difference is apparent 
even after several months. Ten per cent, of carboxyhaemo- 
globin can be detected in blood by tests 4 and 5. 

6. Copper Sulphate Test — A drop of saturated copper sul- 
phate solution added to 2 cc. of carbon monoxide blood mixed 
with the same volume of water gives a brick-red precipitate. 
The deposit from normal blood is greenish brown. In all these 
precipitation tests (4, 5 and 6) the less easily decomposed 
carbon monoxide blood remains bright red but the more easily 
decomposed normal blood in presence of the precipitants used 
and others is off color or dark. 

7. Ammonium Sulphide Test. — Mix 0.2 cc. of ammonium 
sulphide solution and 0.2-0.3 cc. of 30 per cent, acetic acid with 
10 cc. of 2 per cent, aqueous blood solution. Carbon monoxide 
blood gives a fine rose color but normal blood is greenish gray. 
The former within 24 hours gives a red flocculent precipitate. 

8. Palladous Chloride Test — Carbon monoxide precipitates 
black metallic palladium from a neutral aqueous palladous 
chloride solution: 

CO -f H2O -h PdCl, = CO, -h 2HCI -f- Pd. 

^ Mix I volume of glacial acetic acid with 2 volumes of water. This acid con- 
tains about 30 per cent, acetic acid. 
* Use I part of blood to 4 parts of water. 



DETECTION OF CAHBON MONOXIDE BLOOD 



299 



Mix a few drops of potassium hydroxide solution with the blood 
and warm gently upon the water-bath. By means ot a suction 
pump draw through the solution air that has been washed until 
pure. Pass the gas evolved first through lead acetate solution 
to remove possible hydrogen sulphide, then through sulphuric 
add to absorb ammonia and finally through a neutral light red 
palladous chloride solution (i ooo). 

9. Spectroscopic Ezaminatioii. — The detection of carboxy- 
hEemoglobin with ;the spectroscope is comparatively easy. The 






m 



Carbo lyhaemoglobiD. 



Haem&Ioiiorphy 



HaemmtoporphyriQ, 



I F[C. 34.— Absorption -Spectra. 

two absorption bands of this compound are quite similar to 
those of oxyhemoglobin but they lie somewhat nearer together 
and more toward the violet. The main difference, however, 
between the absorption bands of these compounds is that those 
of car boxy hipmoglobin are not extinguished by reducing agents. 
To prepare the blood solution for spectroscopic examination, 
dilute 1-1.5 parts of blood with 100 parts of water and make the 



300 DETECTION OF POISONS 

observations through a layer i cm. thick. To reduce i per 
cent, blood solution, mix thoroughly with a few drops of am- 
monium sulphide solution and add 4-6 drops more of the same 
reagent as a surface layer to exclude air. Reduction begins in 
about 6-8 minutes. A solution of tartaric add and ferrous 
sulphate in presence of an excess of ammonium hydroxide 
solution will also reduce oxyha^moglobin. 

Oxyhaemoglobin under these conditions is changed to reduced 
haemoglobin. The two absorption bands characteristic of the 
former disappear and a broad diffuse absorption band occupies 
the previous bright space between the two bands. The spectrum 
of carboxyhajmoglobin remains unchanged only when 27 per 
cent, at least of the haemoglobin is saturated with carbon 
monoxide. If allowed to stand in an open vessel, blood will lose 
carbon monoxide within 8 days. But carbon monoxide blood 
sealed in glass tubes is said to keep for years. Carbon 
monoxide has been detected in blood of a cadaver after 18 
months. 

Tollens^ recommends adding some formaldehyde to the blood solution. This 
reagent has not the slightest efifect upon the two oxyhemoglobin bands. Warm- 
ing the mixture very gently with ammonium sulphide solution develops a third 
and nearly as distinct black band almost midway between the original bands 
which gradually disappear. Finally only this band will remain. This is a far 
more satisfactory test than that given by the indefinite band of blood alone. 
If the solution is cooled and agitated with air, this third band will disappear and 
the two original oxyha;moglobin bands will return. 

If carbon monoxide is present, formaldehyde does not have this action. 

2. Detection of Blood Stains 

The detection of blood in dry stains upon fabrics, wood, knives, 
weapons, etc.,- is more certain and less open to question, if 
hsemin crystals (Teichmann's blood crystals) are prepared from 
the blood pigment. If haemin crystals are obtained, the stain 
in question may be regarded w^ith certainty as due to blood. 
Fresh blood when dry is bright red and has a smooth surface. 

* Berichte der Deutschen chcmischen Gesellschaft 34, 1426 (1901). 

* Blood mixed with iron oxide as, for example, blood upon rusty knives and 
weapons usually fails to give haemin crystals. 



DETECTION OF CARBON MONOXIDE BLOOD 



301 



Flakes of such blood scraped from any material are garnet red 
by transmitted light. A solution of fresh blood stains in potas- 
sium or sodium hydroxide is dichroic, being red by transmitted 
^-Biid green by reflected light. Later dried blood becomes 
Pbiownish red or dark brown. These color changes are due to 
' conversion of oxyhemoglobin into methasmoglobin and then 
into hiematin. The first two substances are soluble in water 
but the last is not. But hsematin is soluble in alkalies and in 
aicohol containing sulphuric acid. This change of the blood 
pigment depends not only upon the age of the stain but really 
upon the action of air (oxygen), light, heat and moisture upon 
the blood before it is dry. If the blood is in a thin layer, 
haemoglobin will sometimes change into methsmoglobin even 
in 3-10 days. Boiling water causes immediate insolubility. 
The action is also very rapid in direct sunlight. Washing in 
alkaline solutions (boiling solutions of potassium or sodium 
soap, sodium carbonate solution, ammonia and sewage) also 
causes rapid decomposition. But acids, nitric and hydrochloric, 
as well as putrefaction, act more slowly, giving the blood a 
laked appearance and even making it clear and colorless. If, 
however, the blood has once dried, iJiese injurious agencies, 
even putrefaction, act less easily. 

Preparation of Haemin Crystals. — Prepare a cold aqueous 
extract of the stain as free as possible from libers and evaporate 
the solution upon a watch glass away from dust. Add a trace 
of sodium chloride' to the residue, also 8-10 drops of glacial 
acetic acid, and stir with a glass rod. Heat just for an instant 
over a small flame, then evaporate the solution gradually 
upoh a moderately warm water-bath and examine the residue 
with a microscope magnifying 300-500 times. If hfemin 

' Strzyzowski (Chemisches CentralbUtt, 1897, I, ags) advises using sodium 
iodide instead of sodium chloride. Place a small particle oF material suspected 
of containiog blood upon a glass slide and add a drop of sodium iodide solutioQ 
(i ; soo). Evaporate and cover with a cover-glass. Heat for 3-6 seconds 
with concentrated acetic acid which is allowed to run under the cover-glass. The 
test with this modification is said to be more delicate, owittg to the darker color 
of the hiemalin bydriodide co'S'als. The cr>'stals are usually obtained in less 
time and with as small a quantity as o.oooojs gram of fresh blood. Tr. 




302 DETECTION OF POISONS 

cr3^tals fail to appear, repeat the evaporation several times, 
using in each instance 8~io drops of glacial acetic add, and 
examine the residue each time under the microscope. Haemin 
crjrstals are brownish red to dark brown and form rhombic scales 
which frequently lie crossed (Fig. 25). Usually glacial acetic 

add is the only solvent that will ex- 
tract the pigment from old blood 
stains. Brucke heats the stains or 
scrapings to boiling in a test-tube 
with 10-20 drops of gladal acetic 
acid. The decanted or filtered solu- 
tion, after addition of a trace of 
sodium chloride, is evaporated upon 
a watch glass to dryness at 40-80° 
Fig. 25.— H«min Crystals. and the residue is examined under 

the microscope. By this method it 
is immaterial whether the blood has coagulated or not. 

Cold water is without effect upon blood stains, if they have 
previously been treated with hot water. Protein substances in 
the blood are thus coagulated and rendered insoluble. In 
such a case treat the stain with water containing a few drops of 
sodium hydroxide solution. If the stains are upon wool, use 
very dilute sodium hydroxide solution since alkalies dissolve 
wool. Water containing ammonium hydroxide will extract 
stains and this alkali does not act upon wool. Use the alkaline 
aqueous extract to prepare haemin crystals. Evaporate the 
solution to dryness in a watch glass upon the water-bath and 
mix the residue intimately with 8-10 drops of gladal acetic add. 
Add a trace of sodium chloride and again evaporate. Sometimes 
it is advisable, after addifying the extract of the stain with 
acetic acid, to add tannic add, or zinc acetate, and prepare 
Teichmann's crystals from the predpitate. 

Occasionally it is necessary to extract suspected stains with 
hot alcohol containing sulphuric add. Haematin formed from 
the blood pigment dissolves. If this compound is present, the 
solution has a brown color. Excess of sodium hydroxide solu- 
tion will produce the dichroism characteristic of an alkaline 



I 



I 



I 



I DETECTION OF CARBON MONOXIDE BLOOD 303 

hffimatm solution, namely, red by transmitted and green by 
reflected light. Obviously, hiematin should be identified by 
the spectroscope both in acid and alkaline solution. 

Blood mixed with iron oxide (blood upon rusty weapons) 
usually fails to give hiemin crystals but the extract with dilute 
sodium hydroxide solution frequently shows the dichroism of 
luematin solution. Since iron oxide or rust forms an insoluble 
compound with hasmatin, warm such stains for some time upon 
the water-bath with sodium hydroxide solution to dissolve any 
hjematin present. 

HmnBtiii.—Warmine an aqueous blood solution to about 70° decomposes the 
blood pigment oxyhemoglobin into a protein substance called globin and hiematio 
a pigment containing iron. Adda, alkalies and several metallic salts decompose 
oxyhemoglobin in the same way. If this decomposition takes place in the ab- 
sence of oxygen, another pigment appears. Hoppe-Seyler gave the latter the 
name hiemochramogen and other experimenters have called it "reduced hxmatin." 
Oxygen and consequently air rapidly oxidizes this pigment to hiemalin. On the 
other hand reducing agents like ammonium sulphide convert himatinintohsmo- 
chiotnogen. Different formulas are given for hiematin. W. Kflster and others 
now give it the formula CiiHiiNiFeO|. H«ematin Is amorphous and has a dark 
brown or blue-black color. In water, dilute acids, alcohol, ether and chloroform 
it is insoluble but soluble in alcohol or ether containing acid. In even very dilute 
solutions of caustic alkalies it is freely soluble. Alkaline hxmatin solutions are 
dichroic. In rather thick layers the color appears red by transmitted light and 
greenish in thin layers. Add solutions are always brown. Alkaline luematin 
solutions are precipitated by calcium or barium hydroxide solution. 

TTfBTTiiTi is the hydrochloric ester of hsmatin. Very prob- 
ably b^min has the empirical formula C34HaaN4Fe04Cl. 

Note.— If the blood stain is perfectly fresh, it may be recog- 
nized by observing blood-corpuscles with the microscope. 
Human blood can be differentiated from animal blood by com- 
paring blood-corpuscles with those of animal blood as to size, 
only when the corpuscles are still intact. 

Spectroscopic Detectioa of Blood 

" If the extract of a blood stain with cold water is already 
brown, a third fainter and narrower band will appear in ad- 
dition to the two oxyhsemoglobin bands. This lies in the 
orange between C and D and is the methsemoglobin band. 



304 DETECTION OF POISONS 

Cold water will dissolve most of the methaemoglobin from 
fresh, dried blood stains. 

Acetic acid will discharge these two bands, if the oxyhemo- 
globin solution is not too dilute. At the same time the solution 
will become mahogany-brown from formation of hsematin in 
acid solution. This solution has a characteristic spectrum, 
namely, four absorption bands in the yellow and green. If 
excess of ammonium hydroxide is added to this solution, the 
alkaline solution contains haematin, recognizable by a broad 
faint absorption band lying between the red and yellow. A 
few drops of ammonium sulphide solution will extinguish this 
band and bring out two broad bands, namely, one in the green 
and the other in the light blue. These bands lie farther to the 
right than do those of oxyhaemoglobin and are of about the 
same width. This is the spectrum of reduced haematin (haemo- 
chromogen). All these spectroscopic tests are very charac- 
teristic, especially the spectra of oxyhaemoglobin, haemoglobin 
and, in the case of old blood stains, that of reduced haematin. 

Up to the present time no red solution has been found which, 
upon abstraction and addition of oxygen, will give the same 
spectroscopic phenomena as blood. 

When the quantity of blood is very small, or when the blood 
pigment has undergone further decomposition, so that the 
bands of oxyhaemoglobin are no longer visible, it is advisable 
to extract the stains for several hours with concentrated potas- 
sium cyanide solution. Blood will give a light red or yellowish 
brown solution containing the cyano-compound of haematin. 
The spectrum of haematin in alkaline solution will appear as a 
broad, faint band. 

The investigations of Kratter and Hammerl have shown that 
charred blood, which no longer responds to any of the other 
blood reactions, will still give the haematoporphyrin spectrum 
upon treatment with concentrated sulphuric acid (E. v. Hof- 
mann, Lehrbuch der gerichtlichen Medizin, 1903). 

Ammoniacal carmine solution gives two absorption bands 
similar to those of oxyhaemoglobin but they do not change 
upon addition of acetic acid or ammonium sulphide. A band 



DETECTION OF CARBON MONOXIDE BLOOD 305 

given by fuchsine analogous to that of haemoglobin remains 
unchanged after agitation with air. 

Other Blood Tests 

I. Sch5nbein-Van Deen Ozone Test. — A mixture of ozon- 
ized turpentine^ and alcoholic tincture of giiaiac resin, shaken 
with a little blood, produce a light blue color. Separated from 
the turpentine, the tincture is deep blue. Though very 
delicate, this test is not characteristic of blood, for many 
inorganic and organic substances under the same conditions 
produce ''guaiac blue." Nitrous acid, chlorine, bromine and 
iodine, chromic and permanganic acids, ferric and cupric salts 
produce blue solutions direct with guaiac resin. In examining 
blood stains usually it is possible to exclude these substances 
beforehand. But other substances like cell contents or haemo- 
globin, having the power of transferring ozone, may attach 
false significance to the guaiac-blue reaction. Enzymes 
(diastases), hydrolytic ferments (enzymes in the narrower 
sense), as well as the so-called oxidation ferments (oxidases), are 
organic substances of this character. They occur in different 
parts of plants, especially in fungi and in seeds. Saliva, ex- 
tracts of certain organs, contents of white blood-corpuscles and 
pus cells are animal products of similar nature. E. Schaer^ 
states that these animal and vegetable substances differ from 
hydrogen dioxide in being catalytic in action and carriers of oxy- 
gen at the same time. And also that a temperature of loo®, 
or contact with hydrocyanic acid, completely destroys their 
power of transferring oxygen, or at least greatly diminishes it, in 
which respect they are essentially different from haemoglobin. 
Neither high temperature (ioo°) nor hydrocyanic acid has any 
restraining influence upon haemoglobin so far as transference of 
oxygen is concerned. Consequently an extract, containing one 
of these ferment-like substances but no blood, placed even for a 

^ Turpentine always contains ozone, if exposed to light for a long time in a 
loosely stoppered bottle. 

* Forschungsberichte Uber Nahrungsmittel, etc., 3, i (1896) and Archiv der 
Pharmazie 236, 571 (1898). 
20 



306 DETECTION OF POISONS 

short time in a hot water-bath, loses the power of giving the 
"guaiac blue'' test. In absence of blood, the result will also 
be negative, if the extract of the suspected stain is treated with 
hydrocyanic acid. For these reasons great care is necessary in 
interpreting a positive guaiac test given by the extract of a 
supposed stain. The guaiac test is certainly very usefid as a 
delicate preliminary test and in many instances as a check upon 
biood. The three forms of the blood pigment entering into 
such an examination, namely, haemoglobin, methsemoglobin 
and haematin, are alike in the guaiac test, at least qualitatively, 
as far as transference of oxygen is concerned. The examination 
and extraction of the stain may, therefore, be conducted in 
neutral, acid or alkaline solution, depending upon the nature of 
the substance, and either hot or cold. Render an alkaline 
extract faintly acid with acetic acid before adding guaiac tinc- 
ture. In many instances it is advisable to extract the blood 
stain with hot alcohol containing sulphuric acid. Treat such an 
acid, alcoholic haematin solution with guaiac tincture direct. 
Addition of water will precipitate the resin with the adherent 
blood pigment. 

(a) Vitali's Procedure. — Extract the stain with water con- 
taining carbon dioxide, or old stains with very dilute sodium 
hydroxide^ solution free from nitrite and nitrate. Filter the 
extract and add a little alcoholic guaiac tincture to a portion of 
the filtrate after acidification with acetic acid, if necessary. 
If the milky liquid is not blue in 15 minutes, interfering oxidizing 
agents are absent. Then add a few drops of old turpentine and 
shake. The milky liquid will turn blue at once, or in a short 
time, if blood pigment is present. Very gentle warming upon 
the water-bath increases the delicacy of the reaction. * Even 
putrid blood 2 months old is said to give a positive test. 

(b) E. Schaer's Procedure. — Blood stains upon linen, though 
quite old, dissolve completely when treated for some time with 
70 per cent, chloral hydrate solution. Moistening the stains 
beforehand with glacial acetic acid aids solution. Also pre- 
pare an extract of guaiac resin in 70 per cent, chloral hydrate 

^ Use sodium hydroxide prepared from metallic sodium in this test. 



DETECTION OF CARBON MONOXIDE BLOOD 307 

solution. Mix the extract of the stain with an equal volume of 
the latter solution. In absence of nitrites, the color of ^is 
mixture is brownish yellow to light brown. If preferreoj^m^ 
contact test for blood may be made by this method. Add to 
the mixture of blood and guaiac Hiinefeld's^ turpentine solu- 
tion, or hydrogen peroxide, as a surface-layer. An intense 
blue zone will appear where the two solutions meet. Guaiaconic 
acid in guaiac resin produces "guaiac blue." O. Dobner has 
suggested substituting a dilute solution of guaiaconic acid for 
guaiac resin. Blood, or blood pigment, behaves like a ferment 
and activates the ozonized turpentine or hydrogen peroxide, 
either of which by itself will not turn the solution of guaiac 
resin blue. 

2. Schaer's Aloin Test — The same conditions, producing 
"guaiac blue" from guaiaconic acid, give rise to "aloin red" 
from aloin. This substance has a stronger coloring power and 
lasts longer than " guaiac blue." Use the same solution of blood 
in 70-75 per cent, chloral hydrate solution mixed with a weak 
chloral hydrate solution of aloin. Add Hiinefeld's hydrogen 
peroxide solution as a surface layer. After some time a violet- 
red zone will appear and a red color of equal intensity will 
gradually extend throughout the aloin solution. Another 
method of making this test consists in first extracting the blood 
stain with pure water, acetic acid, chloral hydrate solution or 
alkaline salt solution. Neutralize this solution and add dilute 
alcoholic aloin solution and hydrogen peroxide. If the sus- 
pected stain contains blood pigment, a red color will appear at 
once and persist for a long time. 

3. Biological Detection of Human Blood' 

Injection of bacteria produces specific, bacteriolytic bodies 
and similarly injection of the blood of one animal species into 

* See page 314 for the preparation of this reagent. 

* This subject has been introduced for the sake of completeness. If such an 
investigation is for forensic purposes, the chemist will either decline to undertake 
it, or conduct the experiment with an associate who has had bacteriological and 
pathological experience. 



308 DETECTION OF POISONS 

an animal of a different species gives rise to specific, haemolytic 
and agglutinating bodies. Rabbit's blood, for example, in- 
jected repeatedly into a guinea pig, develops in the serum of 
such a guinea pig substances capable of agglutinating and dis- 
solving red corpuscles of the rabbit, setting haemoglobin free 
and rendering the blood laky. Blood serum from an animal, 
into which defibrinated blood, or blood serum from a different 
animal species has been injected intravenously, subcutaneously 
or intraperitoneally, that is to say, into the peritoneal cavity, 
has the peculiar property of causing precipitation only in blood 
serum of this particular animal species. Uhlenhuth,^ Wasser- 
mann and Schiitze,^ and others have made independent experi- 
ments of this kind with blood serum to find for forensic purposes 
a test, based upon this biological method, which shall differenti- 
ate human blood from the blood of every other animal species. 
Repeated injection of lo cc. of defibrinated human blood, or 
human blood serum free from cells, into a rabbit, either intra- 
peritoneally or subcutaneously, yields a serum producing a 
heavy, cloudy precipitate in an aqueous solution of human blood. 
This coagulin is specific in action, producing a precipitate only 
in presence of human blood. Wassermann and Schutze tested 
the blood of 23 different animals, among which were mammals, 
birds and fishes, and obtained negative results with blood 
solutions from these very different animal species. By use of 
blood serum it is possible to differentiate even old human blood, 
dried for many weeks, from the blood of other animals. 

To demonstrate the use of this method, A. Dieudonn6' 
prepares i per cent, blood solutions, placing 2 cc. of the clear 
filtered solution in small test-tubes and adding an equal volume 
of double physiological salt solution (= 1.8 per cent. NaCl). 
Then add 6 drops of serum to each portion and place the tubes 
in an incubator at 37°. The serum of the rabbit, treated with 
human blood serum, added to an aqueous solution of human 

1 Deutsche medizinische Wochenschrift, 1901, No. 6; und Zeitschrift fOr 
Medizinalbeamte, 1903, Heft 5 and 6. 

* Berliner klinische Wochenschrift, 1901, No. 7. 
•Munchener medizinische Wochenschrift, 1901, page 533. 



DETECTION OF CARBON MONOXIDE BLOOD 309 

blood, produced in a few minutes a distinct flocculent precipi- 
tate which gradually became more and more marked. As a 
check, test also with normal rabbit's serum which will cause no 
precipitate in a solution of human blood. Dieudonn6 found also 
that rabbit's serum, obtained after injecting human blood serum, 
causes precipitates not only in human blood solutions but in 
human urine containing albumin, with an exudate from human 
pleura and with peritoneal exudate. But precipitation in the 
case of human blood was much more marked than in these other 
tests. In his experiments Dieudonn6 used blood expressed 
from the placenta, repeatedly injecting it subcutaneously into 
rabbits in separate doses of lo cc. and at intervals of 3-4 days. 
The animals were bled several days after the last injection and 
the blood was kept upon ice. 

The antiserum used in detecting blood should above every- 
thing else be perfectly clear. To prepare such serum, use a 
sterile Berkefeld filter attached to a water pump. The anti- 
serum should be active in very dilute solution. Distinct tur- 
bidity should appear immediately in a solution diluted i : 1000, 
or in 1-2 minutes at latest. Sera must be of this high efiiciency 
for practical use. Uhlenhuth has shown that the biological 
method of detecting blood is specific for human albumin. A 
necessary consequence of this fact is that the material should 
first of all be shown to be blood. The first question for the 
expert to answer in such an investigation must always be: 
**Is there any blood at all present?" If the answer is affirma- 
tive, the next question is: "Is it human or animal blood?" 
Consequently the material should first be examined for blood 
stains by van Deen's ozone test, Teichmann's haemin test and 
by the spectroscope. If the suspected stains are upon a hard 
surface, as a knife, hatchet, gun barrel, wood, stone, etc., they 
should be scraped oflf for the biological blood test and extracted 
for several hours in a test-tube with physiological salt solution 
(= 0.9 per cent. NaCl). First, filter the extract through paper. 
If the filtrate is not clear, next use a Berkefeld filter. 



APPENDIX 
PREPARATION OF REAGENTS 

General Alkaloidal Reagents. — ^A class of reagents, known as 
general alkaloidal reagents, added to solutions of most of the 
alkaloids or of their salts, produce precipitates characterized by 
their color, their amorphous or crystalline appearance and their 
insolubility or sparing solubility in water. But these reagents 
do not precipitate alkaloids exclusively. Several members of 
this class, for example, the chlorides of gold, platinum and 
mercury, phospho-molybdic and phospho-tungstic acids, react 
similarly with ammonia and many ammonium derivatives. An 
explanation of this similarity in behavior is found in the fact 
that most of the alkaloids, being secondary or tertiary bases, are 
themselves ammonium derivatives. Nearly all the general 
alkaloidal reagents also precipitate proteins, albumoses, pep- 
tones, creatinine and the nuclein bases, adenine, guanine, hy- 
poxanthine and xanthine. 

The general alkaloidal reagents are especially useful in 
detecting the presence, or absence, of alkaloids and other basic 
compounds. If there is only a slight residue from the ether 
extract of the alkaline solution in the Stas-Otto method, test 
first with the general alkaloidal reagents and then, if neces- 
sary, for individual alkaloids. To perform these tests, dissolve 
the given residue in very dilute hydrochloric or sulphuric acid, 
distribute the filtered solution upon several watch glasses and 
add to each portion a drop of the more sensitive reagents. 
If an alkaloid or any other basic substance is present, distinct 
precipitates or at least decided cloudiness will appear in all or 
in nearly all of the tests. 

The most important general alkaloidal reagents are the 
following: 

Gold Chloride dissolved in water (i :3o) produces white, yel- 
low or brown precipitates which are amorphous or crystal- 

310 



PREPARATION OF REAGENTS 311 

line. These precipitates decompose to some extent with 
separation of metallic gold. 

Platinum Chloride dissolved in water (i : 20) produces yellow- 
ish white to yellow precipitates which are usually granular and 
crystalline. These precipitates are usually analogous in com- 
position to ammonium chloroplatinate, (H4N)2PtCl6. 

Mercuric Chloride dissolved in water (i : 20) produces white 
to yellowish precipitates which are usually amorphous but 
gradually become crystalline. 

lodo-potassiiim Iodide, prepared by dissolving 5 parts of 
iodine and 10 parts of potassium iodide in 100 parts of water, 
produces brown precipitates which are usually flocculent. 

Potassium Cadmium Iodide, prepared by dissolving 20 grams 
of potassium iodide in 20 cc. of boiling water, adding 10 grams 
of cadmium iodide and diluting to 100 cc, produces white or 
yellowish precipitates with sulphuric acid solutions of most of 
the alkaloids, even when these solutions are very dilute. These 
precipitates, at first amorphous but later crystalline, dissolve 
in an excess of the reagent and also in alcohol. 

Potassium Bismuthous Iodide may be prepared according to 
Kraut^ by dissolving 80 grams of bismuth subnitrate in 200 
grams of nitric acid (sp. gr. 1.18 = 30 per cent. HNO3) and pour- 
ing this solution into a concentrated solution of 272 grams of 
potassium iodide in water. Allow the potassium nitrate to 
crystallize and dilute the solution with water to 1000 cc. This 
reagent produces orange-red precipitates with sulphuric add 
solutions of many alkaloids. By shaking these precipitates with 
sodium hydroxide and carbonate solution, it is often possible 
to recover the alkaloids unchanged and sometimes almost 
quantitatively. 

Potassium Merctuic Iodide, prepared by ' dissolving 1.35 
grams of mercuric chloride and 5 grams of potassiimi iodide in 
100 cc. of water, produces white or yellowish precipitates with 
hydrochloric acid solutions of most of the alkaloids. These 
precipitates at first amorphous, gradually become crystalline. 

1 Annalen der Chemie und Pharmazie, 210, 310 (1882) und Archiv der Phar- 
mazie, 235, 152 (1897). 



312 DETECTION OF POISONS 

Potasshim Zinc Iodide is prepared by dissolving lo grams of 
zinc iodide and 20 grams of potassium iodide in 100 cc. of water. 

Phospho-molybdic Acid may be prepared by either of the 
following methods: 

(a) Saturate sodium carbonate solution with pure molybdic 
acid, add i part of crystallized disodium phosphate (NaiHPOi.- 
12H2O) to 5 parts of the acid and evaporate to dryness. Fuse 
the residue in a porcelain crucible and dissolve the cold melt in 
water. Prepare 10 parts of solution from i part of this residue. 
Add enough nitric acid to the filtered solution to produce a 
golden yellow color. 

(b) If molybdic acid is not at hand, completely precipitate at 
40° with excess of sodium phosphate solution the nitric acid 
solution of ammonium molybdate used in testing for phos- 
phoric acid. Thoroughly wash the yellow precipitate, add 
water and dissolve in warm concentrated sodium carbonate 
solution. Evaporate this solution to dryness and fuse the resi- 
due until ammonia is completely expelled. If there is any 
reduction (blue or black color), moisten the residue with nitric 
acid and fuse again. Dissolve this residue in hot water and 
add nitric acid in large excess. Prepare 10 parts of solution 
from I part of residue. The golden yellow solution should be 
protected from ammonia vapor. 

Phospho-molybdic acid produces yellowish, amorphous pre- 
cipitates with sulphuric acid solutions of most of the alkaloids. 
After a while these precipitates are frequently greenish or 
bluish from reduction of molybdic acid to molybdic oxide. 

Phospho-tungstic Acid, prepared by adding a little 20 per 
cent, phosphoric acid to an aqueous solution of sodium tung- 
state, produces precipitates similar to those given by phospho- 
molybdic acid. 

Tannic Acid is a 5 per cent, aqueous solution of tannin. This 
reagent produces whitish or yellowish, flocculent precipitates 
partially soluble in hydrochloric acid. Alkaloids may be 
recovered in part from these precipitates by treating them with 
lead or zinc carbonate, evaporating to dryness and extracting 
the residue with ether, alcohol or chloroform. 



PREPARATION OF REAGENTS 313 

Picric Acid is a concentrated aqueous solution of picric acid 
which produces yellow crystalline precipitates, or amorphous 
precipitates which soon become crystalline. 

Picrolonic Acid is^ used as o.i normal alcoholic solution by 
dissolving 26.4 grams of solid picrolonic acid (CioHgNiOs) in a 
liter of alcohol. With most of the alkaloids this solution pro- 
duces salts called picrolonates which are crystalline, difficultly 
soluble and yellow to red in color. Picrolonic acid behaves to- 
ward bases like a monobasic acid.^ 

B. Other Reagents and Sohitions 

Erdmann's Reagent. — Sulphuric acid containing nitric acid, 
prepared by adding to 20 cc. of pure concentrated sulphuric 
acid 10 drops of a solution of 6 drops of concentrated nitric acid 
in 100 cc. of water. 

Froehde's Reagent. — A solution of molybdic acid in sulphuric 
acid, prepared by dissolving 5 mg. of molybdic acid, or sodium 
molybdate, in i cc. of hot, pure concentrated sulphuric acid. 
This solution, which should be colorless, does not keep long. 

Fehling's Solution. — The two following solutions, which 
should be kept separate, are used in preparing this reagent: 

1. Copper Sulphate Solution.— Dissolve 34.64 grams of pure 
crystallized copper sulphate (CUSO4.SH2O) in sufficient water to 
make 500 cc. 

2. Alkaline Rochelle Salt Solution. — Dissolve 173 grams of 
Rochelle salt (K.Na.C4H406.4H20) and 50 grams of sodium 
hydroxide in hot water and dilute this solution when cold to 
500 cc. 

These two solutions, mixed volume for volume, constitute 
Fehling's solution which should be prepared just before being 
used. Fehling's solution, which has been made up and kept, 
should always be tested before being used. The solution should 
not be used, if it gives a red precipitate of cuprous oxide when 
warmed by itself. 

*L. Knorr, Berichte der Deutschen chemischen Gesellschaft, 30, 914 (1897); 
H. Matthes and O. Rammstedt, 2^tschrift fttr analytische Chemie 46, 565 and 
Archiv der Pharmazie 245, 112 (1907). 



314 DETECTION OF POISONS 

Formaldehyde-su^htiiic Add. — Add 2-3 drops of aqueous 
formaldehyde solution (formalin) to 3 cc. of pure concentrated 
sulphuric acid just before using. 

Glinzbiiig's Reagent^ — Dissolve i part of phlorogludnol and 
I part of vanilline in 30 parts of alcohol. This reagent is used 
to detect free mineral acid, especially hydrochloric add, but it 
does not react with free organic adds. 

Hiinef eld's Solution. — Add 25 cc. of alcohol, 5 cc. of chloro- 
form and 1.5 cc. of galdal acetic add to 15 cc. of old turpentine 
which has been exposed for some time to air and light. The 
turpentine used should not produce a blue color with guaiac 
tincture direct nor with 15 cc. of 3-5 per cent, hydrogen peroxide 
free from add. This solution is used in the detection of blood. 

Iodic Acid Solution. — Prepare a 10 percent, aqueous solution 
of iodic add (HIO3). 

Magnesia Mixture. — Dissolve 1 1 grams of crystallized mag- 
nesium chloride (MgCl2.6H20) and 14 grams of ammonium 
chloride in 130 cc. of water and add 70 grams of ammonium 
hydroxide solution (sp. gr. 0.96 = 10 per cent, of NHa). This 
mixture should be clear. It is used to detect arsenic and 
phosphoric adds. 

Mandelin's Reagent. — Dissolve i part of ammonium meta- 
vanadate (H4N.VO3) in 200 parts of pure concentrated sulphuric 
acid. 

Millon's Reagent. — Dissolve i part of mercury in i part of 
cold fuming nitric add. Dilute with twice the volume of 
water and decant the dear solution after several hours. 

Nessler's Reagent. — Dissolve separately in the cold 3.5 
grams of potassium iodide in 10 cc. of water and 1.7 grams of 
mercuric chloride in 30 cc. of water. Add mercuric chloride 
solution to potassium iodide solution until there is a permanent 
predpitate. Dilute with 20 per cent, sodium hydroxide solution 
until the volume is 100 cc. Add mercuric chloride solution, 
until there is again a permanent predpitate and let the solution 

* It is advisable to prepare this reagent as required. Keep two separate alco- 
holic solutions (i : 15) of phlorogludnol and vanilline and mix volume for volume 
as needed. Tr. 



PREPARATION OF REAGENTS 315 

settle. Decant the clear solution and keep in small bottles in 
the dark. This reagent improves upon standing. 

Mecke's Reagent.' — Dissolve 0.5 gram of selenious acid in 
10 grams of pure concentrated sulphuric acid. 

Stannous Chloride Solution. — Mix 5 parts of crystallized 
stannous chloride with i part of hydrochloric acid and com- 
pletely saturate with dry hydrochloric acid gas. Let this 
solution settle and filter through asbestos. It is a pale, 
yellowish^ refractive liquid (sp. gr. at least 1.9). This solution 
is used to detect arsenic (fiettendorff's Arsenic Test). 

C. The Indicator lodeosine 

lodeosme, or erythrosine, CsoHglfOi, is a tetra-iodo-fluoresceine, formed by 
treating fluoresceine with iodine and having the formula: 

1 \CeH4.C0.0 



The commercial preparation usually contains as impurities small quantities of 
substances almost insoluble in ether. To obtain a pure product,' dissolve com- 
mercial iodeosine in aqueous ether and extract iodeosine from the filtered ether 
solution by means of dilute sodium hydroxide solution. Strong sodium hydrox- 
ide solution, added to this aqueous alkaline solution, precipitates the sodium salt 
of iodeosine. Filter, wash with cold alcohol and crystallize from hot alcohol. 
Well formed, almost rectangular plates having a green color on the surface are 
obtained. Hydrochloric acid precipitates pure iodeosine from the aqueous solu- 
tion of the sodium salt. Pure iodeosine dried at 1 20'' is markedly lighter than the 
commercial preparation. It is almost insoluble in absolute ether, benzene and 
chloroform; more easily soluble in acetone, alcohol and aqueous ether. The tone 
of the purified pigment dissolved in aqueous alkali is yellower than that of the 
crude product. lodeosine is a scarlet crystalline powder which dissolves in alcohol 
with a deep red and in ether with a yellowish red color. lodeosine is said to be 
insoluble in water containing a trace of hydrochloric add. To prepare iodeosine 
solution for use as an indicator, dissolve i gram of the pigment in 500 grams of 
alcohol. 

^ Zeitschrift ftir ofifentliche Chemie 5, 350 (1899). 

* Fr. Mylius and F. Foerster, Berichte der Deutschen chemischen Gesellschaft 
24, 1482 (1891). 



INDEX 



Abrin, 221 

Absorption spectra, 299 
Acetanilide, 68 
Acetone, 51 
Acid, cacodylic, 238 
carbolic, 26 

-, with aniline, 34 



hydrochloric, 176 

hydrocyanic, 19 

— , with potassium ferrocyan- 

ide, 25 
hypophosphorous, 8 
iodic, reagent, 314 
meconic, 205 
nitric, 177 
oxalic, 182 

phospho-molybdic, reagent, 312 
phosphorous, 14 
phosphoric, 7 

phospho-tungstic, reagent, 312 
picric, 65 

, reagent, 313 



picrolonic, reagent, 313 

salicylic, 7i 

, in foods and beverages, 243 



selenious, reagent, 207 

sulphuric, 180 

sulphurous, 181 

tannic, reagent, 312 
Acids, mineral, 175 
Aconitine, estimation in aconite root, 

255 
Alcohol, ethyl, 49 

Alkalies, 186 

Alkaloids, Stas-Otto method, 59 

Aloin, reagent, 307 

Aluminium acetate, reagent, 282 

Ammonia, 185 

Aniline, 44, 89 



Antimony, 157 

, fate, distribution and elimina- 
tion, 168 

, mirror and spot, 153 

, quantitative determination, 234 

Antipyrine, 78, 118 

Apomorphine, 122 

Arrhenal, 239 

Arsenic, Marsh-Berzelius test, 149 

, Bettendorff's test, 155 

, biological test, 235 

, bulb-tube test, 155 

, detection, 149 

, distinction from antimony, 153 

, electrolytic detection, 226, 230 

, fate, distribution and elimina- 
tion, 166 

, Fresenius-v. Babo test, 154 

, Gutzeit test, 156, 233 

, in organic compounds, 238 

, in presence of organic matter, 

226 

, isolation as trichloride, 226 

, minute amounts, 240 

, mirror and spot, 153 

, normal, 167 

Assaying of alkaloids by E. Merck, 295 

Atoxyl, 239 

Atropa belladonna, estimation of alka- 
loids, 293 

Atropine, 100 

, estimation, 293 

Barium, 164 
Benzaldehyde, 53 
Berberine, estimation of, 275 
Bettendorflf*s arsenic test, 155 
Biological arsenic test, 235 
Biological blood test, 307 



317 



318 



INDEX 



Bismuth, i6o 

, fate, distribution, dimination, 

173 
Bitter almond water, 53 

Blondlot-Dusart test for phosphorus, 8 

Blood, biological test, 307 

, carbon monoxide, 397 

, coagulation, 222 

, defibrinated, 222 

, spectroscopic test, 303 

, tests, 305 

Blood-stains, 300 

Brucine, 96 

, estimation in nux vomica, 286 



Cadmiimi, 160 

Ca£feine, 79, 118 

, estimation in coffee, tea and 

cola-nuts, 264 

, estimation in cacao and choco- 
late, 291 

Cantharidin, 196 

, estimation in Spanish flies, 256 

Carbolic add, 26 

Carbon disulphide, 42 

, estimation in air, 48 

Carbon monoxide in blood, 247 

Carboxy-hxmoglobin, absorption-spec- 
trum, 299 

Carmine, absorption-spectrum, 304 

Cephaeline, estimation in ipecac, 271 

Chavicine, 280 

Chloral hydrate, 38 

, as a solvent, 244 

Chloroform, 35 

, estimation in cadavers, 37 

Choline, 204 

Chromium, 162 

, fate, distribution and elimina- 
tion, 170 

Cinchona alkaloids, estimation in bark, 
261 

Cinchonidine, 257 

Cinchonine, 257 

Cocaine, loi 

Codeine, 106 

, estimation as picrolonate, 247 



Colchidn, 64 

, estimation in seed and ooniis» 

262 

Coniine, 8$ 

Copper, 157 

, fate, distribution and elimina- 
tion, 171 

Crotin, 222 

Cytisine, 198 

Destruction of organic matter, 141 

Digitalin, 201 

Digitalis glucosides, 200 

Digitonin, 200 

Digitoxin, 200 

Distillation, for phosphorus, 5 

, for volatile poisons, 18 

Emetine, 270 

Erdmann's reagent, 313 

Ergot, 202 

Ergotinine, estimation, 204 

Eserine, 105 

Ethyl alcohol, 49 

Extract of belladonna, estimation, 248, 

294i 29s 
of cinchona, estimation, 259 

of hyoscyamus, estimation, 294 

of oplimi, estimation, 278 

of nux vomica, estimation, a88 

Fehling's solution, 313 
Formaldehyde-sulphuric add, reagent, 

314 
Fresenius-v. Babo apparatus, 154 

Froehde's reagent, 313 

Fuchsine, absorption spectrum, 305 

General alkaloidal reagents, 310 
Githagin, 216 

Gold chloride, reagent, 310 
Guaiac, chloral hydrate solution, rea- 
gent, 305 
Guaiac-copper paper, 21 
Glinzburg's reagent, 314 
Gutzeit's arsenic test, 156, 233 

Hacmatin, 303 



INDEX 



319 



Hsematoporphyrin, absorption-spec- 
trum, 299 

, in urine, 195 

Hsemin crystab, 301 

Hsemochromogen, absorption - spec 
tnmx, 303 

Haemoglobin, absorption-s p e c t r u m, 
299 

Haemolysis, 216 

, toxicity estimated by, 251 

Homatropine, loi 

Human blood, detection, 305 

HUnefeld's solution, 314 

Hydrastine, 112 

, estimation, 274 

Hydrastinine, 113 

Hydrochloric add, 176 

Hydrocyanic acid, 19 

Hydrogen sulphide, arsenic-free, 145 

Hyoscyamine, 99 

lodeosine, 315 
Iodic add, reagent, 314 
Iodoform, 41 

lodo-potassium iodide, estimation of 
alkaloids, 250 

, reagent, 311 

Ipecac, estimation of alkaloid in, 270 
Isopelletierine, 263 

Lead, 160, 164 

, fate, distribution and elimina- 
tion, 169 
Lead paper, test for phosphorus, 3 

Magnesia mixture, reagent, 7, 314 

Maltol, 244 

Mandelin's reagent, 314 

Marsh-Berzelius apparatus, 151 

Mecke's reagent, 315 

Meconic add, 205 

Meconine, 206 

Mercuric chloride, reagent, 311 

, cyanide, 25 

Mercury, 158 

, fate, distribution and elimina- 
tion, 171 
Metallic poisons, 141 



Metals, distribution and elimination, 

Methemoglobin, absorption-spectrum, 

299 
Milk, salicylic add in, 243 
Millon's reagent, 314 
Mineral adds, 175 
Mitscherlich apparatus, 5 
Morphine, 126 
, estimation, 247, 276 

Narcdne, 131 
Narcotine, 108 
Nessler's reagent, 314 
Nicotine, 86 

, estimation in tobacco, 272 

Nitric add, 177 

Nitrobenzene, 42 

Non- volatile organic poisons, 57 

Opium, 205 
Oxalic add, 182 

Oxyhaemoglobin, absorption-spectrum, 
299 

Papaverine, 208 

Paraxanthine, 293 

Pelletierine, 263 

Phenacetine, 70 

Phenol, 26 

Phospho-molybdic add, reagent, 312 

Phosphorous add, 14 

Phosphorus, 5 

, Blondlot-Dusart test, 8 

, estimation, 15 

, in oib, 14, 224 

, Hilger-Nattermann, 11 

, Mitscherlich, 5 

Phospho-tungstic add, reagent, 312 
Physiological salt solution, 216 

test. for atropine, 100 

for cantharidin, 198 

for cocaine, 105 

for physostigmine, 106 

for strychnine, 95 

Physostigmine, 105 
Picraconitine, 254 



320 



INDEX 



Picric add, 65 

, reagent, 313 

Picrotoxin, 61 
Pilocarpine, 210 

, estimation, 279 

Piperidine, 280 
Piperine, 280 

Platinum chloride, reagent, 311 
Pomegranate bark, alkaloids in, 263 
Potassium bismuthous iodide, esti- 
mation of alkaloids, 248 

, reagent, 311 

cadmium iodide, reagent, 311 

chlorate, 187 

, to destroy organic matter, 141 



mercuric iodide, reagent, 311 

zinc iodide, reagent, 312 

Pseudo-pelletierine, 263 
Psychotrine, 276 
Ptomaines, 212 
Pyramidone, 119 

Quinidine, 257 

Quinine, 114 

, estimation, 251, 261 

Ricin, 221 

Salicylic add, 72 
Santonin, 192 

, estimation in wormseed, 282 

, estimation in troches, 284 

Saponins, 213 

Schaer's blood tests, 306, 308 
Scherer's phosphorus test, 3 
Schlererythrin, 203 
Schonbein-Van Deen blood test, 305 
Selenious add, reagent, 207 
Selenium, biological arsenic test, 236 
Silver, 164 

, fate, distribution and elimina- 
tion, 172 



Solanidine, 218 
Solanine, 217 

, estimation, 284 

Stannous chloride, reagent, 315 
Stas-Otto process, 59 
St>'ptidne, estimation, 246 
Str>xhnine, 92 

, estimation with quinine, 251 

, estimation in nux vomica, 291 

Sulphonal, 193 
S>'nopsis of Group I, 55 

II, 134 

Ill, 164 

Tannic acid, reagent, 312 
Teichmann's crystals, 302 
Tellurium, biological arsenic test, 236 
Thebaine, 220 
Theine (see Caffeine). 
Theobromine, estimation in cacao, 291 
Theophylline, 293 
Tin, 157 

, fate, distribution and elimina- 
tion, 174 
Toxalbumins, 221 
Trional, 196 

Uranium, 173 

Van Deen's blood test, 305 
Veratrine, 89 
Veronal, 75 
Volatile poisons, 3 

Wine, salicylic add in, 243 

Zinc, 161 

, fate, distribution and elimina- 
tion, 173 



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